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Sample records for cell density cultivation

  1. High cell density cultivation of probiotics and lactic acid production.

    Science.gov (United States)

    Schiraldi, Chiara; Adduci, Vincenzo; Valli, Vivien; Maresca, Carmelina; Giuliano, Mariateresa; Lamberti, Monica; Cartenì, Maria; De Rosa, Mario

    2003-04-20

    The commercial interest in functional foods that contain live microorganisms, also named probiotics, is paralleled by the increasing scientific attention to their functionality in the digestive tract. This is especially true of yogurts that contain strains of lactic-acid bacteria of intestinal origin, among these, Lactobacillus delbrueckii ssp. bulgaricus is extensively used in the dairy industry and it has been demonstrated to be a probiotic strain. In this work we describe high cell density cultivations of this microorganism also focusing on the stereospecific production of lactic acid. Key parameters such as medium composition (bactocasitone concentration) and diverse aeration conditions were explored. The results showed that the final concentration of biomass in anaerobic fermentation was lower than the one obtained in microaerophilic conditions, while it gave a very high productivity of lactic acid which was present as a racemic mixture in the permeate. Fermentation experiments carried out with air sparging, even at very low flow-rate, led to the production of the sole L(+) lactic acid giving sevenfold increase in biomass yield in respect to the batch cultivation. Finally, a mathematical model was developed to describe the microfiltration bioprocess applied in this research considering an inhibition kinetic and enucleating a suitable mathematical description for the decrease of the transmembrane flux. Copyright 2003 Wiley Periodicals, Inc.

  2. Vitreoscilla hemoglobin expression in engineered Escherichia coli: improved performance in high cell-density batch cultivations.

    Science.gov (United States)

    Pablos, Tania E; Mora, Eugenio Meza; Le Borgne, Sylvie; Ramírez, Octavio T; Gosset, Guillermo; Lara, Alvaro R

    2011-08-01

    High cell-density cultivations are the preferred system for biomolecules production by Escherichia coli. It has been previously demonstrated that a strain of E. coli with a modified substrate transport system is able to attain high cell densities in batch mode, due to the very low overflow metabolism displayed. The use of elevated amounts of glucose from the beginning of the cultivation, eliminates the existence of substrate gradients due to deficient mixing at large-scale. However, the large amounts of oxygen demanded resulted in microaerobic conditions after some hours of cultivation, even at small-scale. In this work, the effect of expressing the Vitreoscilla hemoglobin (VHb) in the engineered strain during batch cultures using high-glucose concentrations was tested. Together, the expression of VHb and the modified substrate transport system resulted in a 33% increase of biomass production compared to the parental strain (W3110) lacking the VHb in batch cultivations using 25 g/L of glucose. When 50 g/L of glucose were used, expression of VHb in the modified strain led to 11% higher biomass production compared to W3110. The VHb also increased the growth rates of the strains by about 30% in the aerobic phase and more than 200% in the microaerobic phase of batch cultivation. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Enzyme controlled glucose auto-delivery for high cell density cultivations in microplates and shake flasks

    Directory of Open Access Journals (Sweden)

    Casteleijn Marco G

    2008-11-01

    Full Text Available Abstract Background Here we describe a novel cultivation method, called EnBase™, or enzyme-based-substrate-delivery, for the growth of microorganisms in millilitre and sub-millilitre scale which yields 5 to 20 times higher cell densities compared to standard methods. The novel method can be directly applied in microwell plates and shake flasks without any requirements for additional sensors or liquid supply systems. EnBase is therefore readily applicable for many high throughput applications, such as DNA production for genome sequencing, optimisation of protein expression, production of proteins for structural genomics, bioprocess development, and screening of enzyme and metagenomic libraries. Results High cell densities with EnBase are obtained by applying the concept of glucose-limited fed-batch cultivation which is commonly used in industrial processes. The major difference of the novel method is that no external glucose feed is required, but glucose is released into the growth medium by enzymatic degradation of starch. To cope with the high levels of starch necessary for high cell density cultivation, starch is supplied to the growing culture suspension by continuous diffusion from a storage gel. Our results show that the controlled enzyme-based supply of glucose allows a glucose-limited growth to high cell densities of OD600 = 20 to 30 (corresponding to 6 to 9 g l-1 cell dry weight without the external feed of additional compounds in shake flasks and 96-well plates. The final cell density can be further increased by addition of extra nitrogen during the cultivation. Production of a heterologous triosphosphate isomerase in E. coli BL21(DE3 resulted in 10 times higher volumetric product yield and a higher ratio of soluble to insoluble product when compared to the conventional production method. Conclusion The novel EnBase method is robust and simple-to-apply for high cell density cultivation in shake flasks and microwell plates. The

  4. Control of microbial activity by flow injection analysis during high cell density cultivation of Escherichia coli.

    Science.gov (United States)

    Ding, T; Bilitewski, U; Schmid, R D; Korz, D J; Sanders, E A

    1993-01-01

    The application of an automated flow injection analysis (FIA) system for on-line determination of microbial activity, during high cell density cultivations of Escherichia coli is reported. Based on a bioelectrochemical principle, the FIA method used a redox mediator (potassium hexacyanoferrate(III)) to facilitate electron transfer from the microorganisms to an electrochemical detector. Assays were carried out using a new sampling device which provided aseptic operation by use of a valve and chemical sterilisation. No sample dilution or pretreatment was necessary for biomass concentrations up to approx. 40 g l-1. The sample volume was 0.5 ml and the overall analysis time was 5 min. FIA signals were found to correlate well with the oxygen uptake rate (OUR). Changes in metabolic activity due to low substrate levels or high inhibitor concentrations in the cultivation medium became obvious from the FIA signals.

  5. High cell density cultivation of six fungal strains efficient in azo dye bioremediation.

    Science.gov (United States)

    Abd El-Rahim, Wafaa M; Mostafa, Enas M; Moawad, Hassan

    2016-12-01

    This work aims at optimizing the high cell density fungal cultivation for producing large quantities of fungal biomass to be used in azo dye residues bioremediation. In our previous studies the efficacy of using certain fungal strains to decolorize a range of commercial textile dyes of different structures (azo, disazo) were investigated. Several promising fungal strains belonging to Aspergillus tubigenesis, Aspergillus niger, Aspergillus terreus, and Aspergillus fumigates demonstrated high capacity in decolorizing various azo dyes. This study focuses on the high cell density cultivation of the fungal strains identified as potential bioremediation agents. The study includes the optimization of all parameters involved in bioprocess development for high cell density cultivation of six promising fungal strains. The growth of the fungal strains was tested on the sucrose medium in 7 l-fermenter. The growth of these fungal strains having the capacity to accumulate large quantities of biomass was also tested in medium containing molasses as a cheap substrate. The residual molasses, biomass dry weight and protein content of the six fungal strains showed that the strains 20 and 2 were marked by the highest protein content. In this study a comparative analysis between the results of dry weight, residual molasses and protein content of geowth of the strains 20, 5 and 2 under uncontrolled and controlled pH of media in batch fermentation was studied to follow the accumulation of biomass and protein production in the growth media. The results indicate that the dry weight accumulated by strains No. 20, 5 and 2 grown on molasses was better than those of strains grown on sucrose. Fungal strain No. 5 had the highest biomass dry weight accumulation. The study shows that the molasses as cheaper sugar sources were better than sucrose for growing fungal biomass.

  6. High cell density cultivation of six fungal strains efficient in azo dye bioremediation

    Directory of Open Access Journals (Sweden)

    Wafaa M. Abd El-Rahim

    2016-12-01

    Full Text Available This work aims at optimizing the high cell density fungal cultivation for producing large quantities of fungal biomass to be used in azo dye residues bioremediation. In our previous studies the efficacy of using certain fungal strains to decolorize a range of commercial textile dyes of different structures (azo, disazo were investigated. Several promising fungal strains belonging to Aspergillus tubigenesis, Aspergillus niger, Aspergillus terreus, and Aspergillus fumigates demonstrated high capacity in decolorizing various azo dyes. This study focuses on the high cell density cultivation of the fungal strains identified as potential bioremediation agents. The study includes the optimization of all parameters involved in bioprocess development for high cell density cultivation of six promising fungal strains. The growth of the fungal strains was tested on the sucrose medium in 7 l-fermenter. The growth of these fungal strains having the capacity to accumulate large quantities of biomass was also tested in medium containing molasses as a cheap substrate. The residual molasses, biomass dry weight and protein content of the six fungal strains showed that the strains 20 and 2 were marked by the highest protein content. In this study a comparative analysis between the results of dry weight, residual molasses and protein content of geowth of the strains 20, 5 and 2 under uncontrolled and controlled pH of media in batch fermentation was studied to follow the accumulation of biomass and protein production in the growth media. The results indicate that the dry weight accumulated by strains No. 20, 5 and 2 grown on molasses was better than those of strains grown on sucrose. Fungal strain No. 5 had the highest biomass dry weight accumulation. The study shows that the molasses as cheaper sugar sources were better than sucrose for growing fungal biomass.

  7. Increased bacterial cell density and recombinant protein yield using a commercial microbial cultivation system.

    Science.gov (United States)

    Peck, Grantley R; Bowden, Timothy R; Shiell, Brian J; Michalski, Wojtek P

    2014-01-01

    EnBase (BioSilta, Finland) is a microbial cultivation system that replicates fed-batch systems through sustained release of glucose by enzymatic degradation of a polymeric substrate. Achievable bacterial cell densities and recombinant capripoxvirus protein expression levels, solubility, and antigenicity using the EnBase system were assessed. BL21-AI Escherichia coli expressing capripoxvirus proteins achieved up to eightfold higher cell densities when grown in EnBase media compared with standard media. Greater yields of capripoxvirus proteins were attained using EnBase media, either through increases in the amount of expressed protein per cell in conjunction with higher cell density or through the increase in cell density alone. Addition of EnBase booster enhanced protein yield for one of the proteins tested but reduced yield for the other. However, the amount of soluble forms of the capripoxvirus proteins tested was not different from that observed from cultures grown under standard conditions. Purified capripoxvirus proteins expressed using EnBase or standard media were assessed for their performance by enzyme-linked immunosorbent assay (ELISA) and were shown to be equally capable of specifically binding capripoxvirus antibodies.

  8. Production of savinase and population viability of Bacillus clausii during high-cell-density fed-batch cultivations

    DEFF Research Database (Denmark)

    Christiansen, Torben; Michaelsen, S.; Wumpelmann, M.

    2003-01-01

    The growth and product formation of a Savinase-producing Bacillus clausii were investigated in high-cell-density fed-batch cultivations with both linear and exponential feed profiles. The highest specific productivity of Savinase was observed shortly after the end of the initial batch phase for all....... The physiological state of the cells was monitored during the cultivations using a flow cytometry assay based on the permeability of the cell membrane to propidium iodide. In the latter parts of the fed-batch cultures with a linear feed profile, a large portion of the cell population was found to have a permeable...

  9. Feeding strategies enhance high cell density cultivation and protein expression in milliliter scale bioreactors.

    Science.gov (United States)

    Faust, Georg; Janzen, Nils H; Bendig, Christoph; Römer, Lin; Kaufmann, Klaus; Weuster-Botz, Dirk

    2014-10-01

    Miniature bioreactors under parallel fed-batch operations are not only useful screening tools for bioprocess development but also provide a suitable basis for eventual scale-up. In this study, three feeding strategies were investigated: besides the established intermittent feeding by a liquid handler, an optimized microfluidic device and a new enzymatic release system were applied for parallel fed-batch cultivation of Escherichia coli HMS174(DE3) and BL21(DE3) strains in stirred-tank bioreactors on a 10 mL scale. Lower fluctuation in dissolved oxygen (DO) and higher optical densities were measured in fed-batch processes applying the microfluidic device or the enzymatic glucose/fructose release system (conversion of intermittently added sucrose by an invertase), but no difference in dry cell weights (DCW) were observed. With all three feeding strategies high cell densities were realized on a milliliter scale with final optical density measured at 600 nm (OD600 ) of 114-133 and final DCW concentrations of 69-70 g L(-1) . The effect of feeding strategies on the expression of two heterologous proteins was investigated. Whereas no impact was observed on the expression of the spider silk protein eADF4(C16), the fluorescence of enhanced green fluorescence protein (eGFP) was reproducibly lower, if an intermittent glucose feed was applied. Thus, the impact of feeding strategy on expression is strongly dependent on the E. coli strain and/or expressed protein. As a completely continuous feed supply is difficult to realize in miniature bioreactors, the enzymatic release approach from this study can be easily applied in all microfluidic system to reduce fluctuations of glucose supply and DO concentrations. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. High-cell-density fed-batch cultivation of the docosahexaenoic acid producing marine alga Crypthecodinium cohnii.

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    De Swaaf, Martin E; Sijtsma, Lolke; Pronk, Jack T

    2003-03-20

    The heterotrophic marine alga Crypthecodinium cohnii is known to produce docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications, during batch cultivation on complex media containing sea salt, yeast extract, and glucose. In the present study, fed-batch cultivation was studied as an alternative fermentation strategy for DHA production. Glucose and acetic acid were compared as carbon sources. For both substrates, the feed rate was adapted to the maximum specific consumption rate of C. cohnii. In glucose-grown cultures, this was done by maintaining a significant glucose concentration (between 5 and 20 g/L) throughout fermentation. In acetic acid-grown cultures, the medium feed was automatically controlled via the culture pH. A feed consisting of acetic acid (50% w/w) resulted in a higher overall volumetric productivity of DHA (r(DHA)) than a feed consisting of 50% (w/v) glucose (38 and 14 mg/L/h, respectively). The r(DHA) was further increased to 48 mg/L/h using a feed consisting of pure acetic acid. The latter fermentation strategy resulted in final concentrations of 109 g/L dry biomass, 61 g/L lipid, and 19 g/L DHA. These are the highest biomass, lipid, and DHA concentrations reported to date for a heterotrophic alga. Vigorous mixing was required to sustain aerobic conditions during high-cell-density cultivation. This was complicated by culture viscosity, which resulted from the production of viscous extracellular polysaccharides. These may present a problem for large-scale industrial production of DHA. Addition of a commercial polysaccharide-hydrolase preparation could decrease the viscosity of the culture and the required stirring. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 666-672, 2003.

  11. Expression of recombinant Pseudomonas stutzeri di-heme cytochrome c(4) by high-cell-density fed-batch cultivation of Pseudomonas putida

    DEFF Research Database (Denmark)

    Thuesen, Marianne Hallberg; Nørgaard, Allan; Hansen, Anne Merete

    2003-01-01

    The gene of the di-heme protein cytochrome c(4) from Pseudomonas stutzeri was expressed in Pseudomonas putida. High-yield expression of the protein was achieved by high-cell-density fed-batch cultivation using an exponential glucose feeding strategy. The recombinant cytochrome c(4) protein...

  12. The effect of intracellular amino acids on GSH production by high-cell-density cultivation of Saccharomyces cerevisiae.

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    Wang, Miaomiao; Sun, Jingfeng; Xue, Feiyan; Shang, Fei; Wang, Zheng; Tan, Tianwei

    2012-09-01

    The present paper studies the effects of precursor amino acids, i.e., L-glutamic acid (Glu), L-glycine (Gly), and L-cysteine (Cys), on the glutathione (GSH) production. The three amino acids were added during the fermentations. The GSH production was analyzed by gas chromatography-mass spectrometry (GC-MS). It was observed that the cell content of Cys reduced continually, Gly maintained a fairly constant concentration, while Glu remained at a high concentration compared with Cys and Gly. The synthesis of GSH was found to significantly increase after 28 h of fermentation upon addition of 6 mmol l(-1) of each of the three amino acids. Under these conditions, the GSH yields reached 2,250±50 mg l(-1) at 34 h from 1,050±50 mg l(-1) at 28 h. The GC-MS analyses on the effect caused by the addition of amino acids indicated that the addition of Glu was not necessary to improve the GSH production by high-cell-density cultivation of Saccharomyces cerevisiae. The addition of Cys or Gly individually enhances the production of GSH.

  13. Bioreactors for high cell density and continuous multi-stage cultivations: options for process intensification in cell culture-based viral vaccine production.

    Science.gov (United States)

    Tapia, Felipe; Vázquez-Ramírez, Daniel; Genzel, Yvonne; Reichl, Udo

    2016-03-01

    With an increasing demand for efficacious, safe, and affordable vaccines for human and animal use, process intensification in cell culture-based viral vaccine production demands advanced process strategies to overcome the limitations of conventional batch cultivations. However, the use of fed-batch, perfusion, or continuous modes to drive processes at high cell density (HCD) and overextended operating times has so far been little explored in large-scale viral vaccine manufacturing. Also, possible reductions in cell-specific virus yields for HCD cultivations have been reported frequently. Taking into account that vaccine production is one of the most heavily regulated industries in the pharmaceutical sector with tough margins to meet, it is understandable that process intensification is being considered by both academia and industry as a next step toward more efficient viral vaccine production processes only recently. Compared to conventional batch processes, fed-batch and perfusion strategies could result in ten to a hundred times higher product yields. Both cultivation strategies can be implemented to achieve cell concentrations exceeding 10(7) cells/mL or even 10(8) cells/mL, while keeping low levels of metabolites that potentially inhibit cell growth and virus replication. The trend towards HCD processes is supported by development of GMP-compliant cultivation platforms, i.e., acoustic settlers, hollow fiber bioreactors, and hollow fiber-based perfusion systems including tangential flow filtration (TFF) or alternating tangential flow (ATF) technologies. In this review, these process modes are discussed in detail and compared with conventional batch processes based on productivity indicators such as space-time yield, cell concentration, and product titers. In addition, options for the production of viral vaccines in continuous multi-stage bioreactors such as two- and three-stage systems are addressed. While such systems have shown similar virus titers compared to

  14. A novel fed-batch based cultivation method provides high cell-density and improves yield of soluble recombinant proteins in shaken cultures

    Science.gov (United States)

    2010-01-01

    Background Cultivations for recombinant protein production in shake flasks should provide high cell densities, high protein productivity per cell and good protein quality. The methods described in laboratory handbooks often fail to reach these goals due to oxygen depletion, lack of pH control and the necessity to use low induction cell densities. In this article we describe the impact of a novel enzymatically controlled fed-batch cultivation technology on recombinant protein production in Escherichia coli in simple shaken cultures. Results The enzymatic glucose release system together with a well-balanced combination of mineral salts and complex medium additives provided high cell densities, high protein yields and a considerably improved proportion of soluble proteins in harvested cells. The cultivation method consists of three steps: 1) controlled growth by glucose-limited fed-batch to OD600 ~10, 2) addition of growth boosters together with an inducer providing efficient protein synthesis within a 3 to 6 hours period, and 3) a slow growth period (16 to 21 hours) during which the recombinant protein is slowly synthesized and folded. Cell densities corresponding to 10 to 15 g l-1 cell dry weight could be achieved with the developed technique. In comparison to standard cultures in LB, Terrific Broth and mineral salt medium, we typically achieved over 10-fold higher volumetric yields of soluble recombinant proteins. Conclusions We have demonstrated that by applying the novel EnBase® Flo cultivation system in shaken cultures high cell densities can be obtained without impairing the productivity per cell. Especially the yield of soluble (correctly folded) proteins was significantly improved in comparison to commonly used LB, Terrific Broth or mineral salt media. This improvement is thought to result from a well controlled physiological state during the whole process. The higher volumetric yields enable the use of lower culture volumes and can thus significantly reduce

  15. High cell density lipid rich cultivation of a novel microalgal isolate Chlorella sorokiniana FC6 IITG in a single-stage fed-batch mode under mixotrophic condition.

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    Kumar, Vikram; Muthuraj, Muthusivaramapandian; Palabhanvi, Basavaraj; Ghoshal, Aloke Kumar; Das, Debasish

    2014-10-01

    A single-stage mixotrophic cultivation strategy was developed with a novel microalgal isolate Chlorella sorokiniana FC6 IITG for high cell density lipid-rich biomass generation. The strain was evaluated for growth and lipid content under different physico-chemical parameters, nutritional conditions and trophic modes. Finally, a single-stage mixotrophic fed-batch cultivation strategy was demonstrated with intermittent feeding of key nutrients along with dynamic increase in light intensity for high cell density biomass and sodium acetate as elicitor for lipid enrichment. The key findings: (i) glucose and sodium acetate was identified as growth supporting and lipid inducing nutrients, respectively; (ii) mixotrophic batch cultivation resulted in maximum biomass and lipid productivity (mgL(-1)day(-1)) of 455.5 and 111.85, respectively; (iii) single-stage mixotrophic fed-batch cultivation showed maximum biomass productivity of 1.93gL(-1)day(-1) (biomass titer 15.81gL(-1)) and lipid productivity of 550mgL(-1)day(-1); (iv) biodiesel properties were in accordance with international standards. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. High-cell-density cultivation of recombinant Escherichia coli, purification and characterization of a self-sufficient biosynthetic octane ω-hydroxylase.

    Science.gov (United States)

    Bordeaux, Mélanie; de Girval, Diane; Rullaud, Robin; Subileau, Maeva; Dubreucq, Eric; Drone, Jullien

    2014-01-01

    We have recently described the biocatalytic characterization of a self-sufficent biosynthetic alkane hydroxylase based on CYP153A13a from Alcanivorax borkumensis SK2 (thereafter A13-Red). Despite remarkable regio- and chemo-selectivity, A13-Red suffers of a difficult-to-reproduce expression and moderate operational stability. In this study, we focused our efforts on the production of A13-Red using high-cell-density cultivation (HCDC) of recombinant Escherichia coli. We achieved 455 mg (5,000 nmol) of functional enzyme per liter of culture. Tight control of cultivation parameters rendered the whole process highly reproducible compared with flask cultivations. We optimized the purification of the biocatalyst that can be performed in either two or three steps depending on the application needed to afford A13-Red up to 95 % homogeneous. We investigated different reaction conditions and found that the total turnover numbers of A13-Red during the in vitro hydroxylation of n-octane could reach up to 3,250 to produce 1-octanol (1.6 mM) over a period of 78 h.

  17. High-cell-density fed-batch cultivation of the docosahexaenoic acid producing marine alga Crypthecodinium cohnii

    NARCIS (Netherlands)

    Swaaf, de M.E.; Sijtsma, L.; Pronk, J.T.

    2003-01-01

    The heterotrophic marine alga Crypthecodinium cohnii is known to produce docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications, during batch cultivation on complex media containing sea salt, yeast extract, and glucose. In the present study, fed-batch

  18. Production of poly(D-3-hydroxybutyrate) from CO(2), H(2), and O(2) by high cell density autotrophic cultivation of Alcaligenes eutrophus.

    Science.gov (United States)

    Tanaka, K; Ishizaki, A; Kanamaru, T; Kawano, T

    1995-02-05

    Hydrogen-oxidizing bacterium, Alcaligenes eutrophus autotrophically produces biodegradable plastic material, poly(D-3-hydroxybutyrate), P(3HB), from carbon dioxide, hydrogen, and oxygen. In autotrophic cultivation of the microorganism, it is essential to eliminate possible occurrence of gas explosions from the fermentation process. We developed a bench-plant scale, recycled-gas, closed-circuit culture system equipped with several safety features to perform autotrophic cultivation of A. eutrophus by maintaining the oxygen concentration in the substrate gas phase below the lower limit for a gas explosion (6.9%). The culture vessel utilized a baskettype agitator, resulting in a K(L) a value of 2970 h(-1). Oxygen gas was also directly fed to the fermentor separately from the other gases. As a result, 91.3 g . dm(-3) of the cells and 61.9 g . dm(-3) of P(3HB) were obtained after 40 h of cultivation under this oxygen-limited condition. The results compared favorably with those reported for mass production of P(3HB) by heterotrophic fermentation. (c) 1995 John Wiley & Sons, Inc.

  19. High cell density media for Escherichia coli are generally designed for aerobic cultivations – consequences for large-scale bioprocesses and shake flask cultures

    Directory of Open Access Journals (Sweden)

    Neubauer Peter

    2008-08-01

    accumulation of formate in oxygen limited cultivations of E. coli can be fully prevented by addition of the trace elements selenium, nickel and molybdenum, necessary for the function of FHL complex. For large-scale cultivations, if glucose gradients are likely, the results from the two-compartment scale-down bioreactor indicate that the addition of the extra trace elements is beneficial. No negative effects on the biomass yield or on any other bioprocess parameters could be observed in cultures with the extra trace elements if the cells were repeatedly exposed to transient oxygen limitation.

  20. High cell density cultivation of Escherichia coli with surface anchored transglucosidase for use as whole-cell biocatalyst for alpha-arbutin synthesis.

    Science.gov (United States)

    Wu, Po-Hung; Nair, Giridhar R; Chu, I-Ming; Wu, Wen-Teng

    2008-02-01

    A fed-batch culture strategy for the production of recombinant Escherichia coli cells anchoring surface-displayed transglucosidase for use as a whole-cell biocatalyst for alpha-arbutin synthesis was developed. Lactose was used as an inducer of the recombinant protein. In fed-batch cultures, dissolved oxygen was used as the feed indicator for glucose, thus accumulation of glucose and acetate that affected the cell growth and recombinant protein production was avoided. Fed-batch fermentation with lactose induction yielded a biomass of 18 g/L, and the cells possessed very high transglucosylation activity. In the synthesis of alpha-arbutin by hydroquinone glucosylation, the whole-cell biocatalysts showed a specific activity of 501 nkat/g cell and produced 21 g/L of arbutin, which corresponded to 76% molar conversion. A sixfold increased productivity of whole cell biocatalysts was obtained in the fed-batch culture with lactose induction, as compared to batch culture induced by IPTG.

  1. Microgravity cultivation of cells and tissues

    Science.gov (United States)

    Freed, L. E.; Pellis, N.; Searby, N.; de Luis, J.; Preda, C.; Bordonaro, J.; Vunjak-Novakovic, G.

    1999-01-01

    In vitro studies of cells and tissues in microgravity, either simulated by cultivation conditions on earth or actual, during spaceflight, are expected to help identify mechanisms underlying gravity sensing and transduction in biological organisms. In this paper, we review rotating bioreactor studies of engineered skeletal and cardiovascular tissues carried out in unit gravity, a four month long cartilage tissue engineering study carried out aboard the Mir Space Station, and the ongoing laboratory development and testing of a system for cell and tissue cultivation aboard the International Space Station.

  2. Development of bioprocess for high density cultivation yield of the probiotic Bacillus coagulans and its spores

    Directory of Open Access Journals (Sweden)

    Kavita R. Pandey

    2016-09-01

    Full Text Available Bacillus coagulans is a spore forming lactic acid bacterium. Spore forming bacteria, have been extensively studied and commercialized as probiotics. Probiotics are produced by fermentation technology. There is a limitation to biomass produced by conventional modes of fermentation. With the great demand generated by range of probiotic products, biomass is becoming very valuable for several pharmaceutical, dairy and probiotic companies. Thus, there is a need to develop high cell density cultivation processes for enhanced biomass accumulation. The bioprocess development was carried out in 6.6 L bench top lab scale fermentor. Four different cultivation strategies were employed to develop a bioprocess for higher growth and sporulation efficiencies of probiotic B. coagulans. Batch fermentation of B. coagulans yielded 18 g L-1 biomass (as against 8.0 g L-1 productivity in shake flask with 60% spore efficiency. Fed-batch cultivation was carried out for glucose, which yielded 25 g L-1 of biomass. C/N ratio was very crucial in achieving higher spore titres. Maximum biomass yield recorded was 30 g L-1, corresponding to 3.8 × 1011 cells mL-1 with 81% of cells in sporulated stage. The yield represents increment of 85 times the productivity and 158 times the spore titres relative to the highest reported values for high density cultivation of B. coagulans.

  3. Biomek Cell Workstation: A Variable System for Automated Cell Cultivation.

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    Lehmann, R; Severitt, J C; Roddelkopf, T; Junginger, S; Thurow, K

    2016-06-01

    Automated cell cultivation is an important tool for simplifying routine laboratory work. Automated methods are independent of skill levels and daily constitution of laboratory staff in combination with a constant quality and performance of the methods. The Biomek Cell Workstation was configured as a flexible and compatible system. The modified Biomek Cell Workstation enables the cultivation of adherent and suspension cells. Until now, no commercially available systems enabled the automated handling of both types of cells in one system. In particular, the automated cultivation of suspension cells in this form has not been published. The cell counts and viabilities were nonsignificantly decreased for cells cultivated in AutoFlasks in automated handling. The proliferation of manual and automated bioscreening by the WST-1 assay showed a nonsignificant lower proliferation of automatically disseminated cells associated with a mostly lower standard error. The disseminated suspension cell lines showed different pronounced proliferations in descending order, starting with Jurkat cells followed by SEM, Molt4, and RS4 cells having the lowest proliferation. In this respect, we successfully disseminated and screened suspension cells in an automated way. The automated cultivation and dissemination of a variety of suspension cells can replace the manual method. © 2015 Society for Laboratory Automation and Screening.

  4. On-line prediction of the feeding phase in high-cell density cultivation of rE. coli using constructive neural networks.

    Science.gov (United States)

    Nicoletti, M C; Bertini, J R; Tanizaki, M M; Zangirolami, T C; Gonçalves, V M; Horta, A C L; Giordano, R C

    2013-07-01

    Streptococcus pneumoniae (pneumococcus) is a bacterium responsible for a wide spectrum of illnesses. The surface of the bacterium consists of three distinctive membranes: plasmatic, cellular and the polysaccharide (PS) capsule. PS capsules may mediate several biological processes, particularly invasive infections of human beings. Prevention against pneumococcal related illnesses can be provided by vaccines. There is a sound investment worldwide in the investigation of a proteic antigen as a possible alternative to pneumococcal vaccines based exclusively on PS. A few proteins which are part of the membrane of the pneumococcus seem to have antigen potential to be part of a vaccine, particularly the PspA. A vital aspect in the production of the intended conjugate pneumococcal vaccine is the efficient production (in industrial scale) of both, the chosen PS serotypes as well as the PspA protein. Growing recombinant Escherichia coli (rE. coli) in high-cell density cultures (HCDC) under a fed-batch regime requires a refined continuous control over various process variables where the on-line prediction of the feeding phase is of particular relevance and one of the focuses of this paper. The viability of an on-line monitoring software system, based on constructive neural networks (CoNN), for automatically detecting the time to start the fed-phase of a HCDC of rE. coli that contains a plasmid used for PspA expression is investigated. The paper describes the data and methodology used for training five different types of CoNNs, four of them suitable for classification tasks and one suitable for regression tasks, aiming at comparatively investigate both approaches. Results of software simulations implementing five CoNN algorithms as well as conventional neural networks (FFNN), decision trees (DT) and support vector machines (SVM) are also presented and discussed. A modified CasCor algorithm, implementing a data softening process, has shown to be an efficient candidate to be

  5. Novel approach of high cell density recombinant bioprocess development: Optimisation and scale-up from microlitre to pilot scales while maintaining the fed-batch cultivation mode of E. coli cultures

    Science.gov (United States)

    2010-01-01

    Background Bioprocess development of recombinant proteins is time consuming and laborious as many factors influence the accumulation of the product in the soluble and active form. Currently, in most cases the developmental line is characterised by a screening stage which is performed under batch conditions followed by the development of the fed-batch process. Performing the screening already under fed-batch conditions would limit the amount of work and guarantee that the selected favoured conditions also work in the production scale. Results Here, for the first time, high throughput multifactorial screening of a cloning library is combined with the fed-batch technique in 96-well plates, and a strategy is directly derived for scaling to bioreactor scale. At the example of a difficult to express protein, an RNase inhibitor, it is demonstrated that screening of various vector constructs and growth conditions can be performed in a coherent line by (i) applying a vector library with promoters and ribosome binding sites of different strength and various fusion partners together with (ii) an early stage use of the fed-batch technology. It is shown that the EnBase® technology provides an easy solution for controlled cultivation conditions in the microwell scale. Additionally the high cell densities obtained provide material for various analyses from the small culture volumes. Crucial factors for a high yield of the target protein in the actual case were (i) the fusion partner, (ii) the use of of a mineral salt medium together with the fed-batch technique, and (iii) the preinduction growth rate. Finally, it is shown that the favorable conditions selected in the microwell plate and shake flask scales also work in the bioreactor. Conclusions Cultivation media and culture conditions have a major impact on the success of a screening procedure. Therefore the application of controlled cultivation conditions is pivotal. The consequent use of fed-batch conditons from the first

  6. High cell density cultivation of Escherichia coli K4 in a microfiltration bioreactor: a step towards improvement of chondroitin precursor production

    Directory of Open Access Journals (Sweden)

    De Rosa Mario

    2011-02-01

    Full Text Available Abstract Background The bacteria Escherichia coli K4 produces a capsular polysaccharide (K4 CPS whose backbone is similar to the non sulphated chondroitin chain. The chondroitin sulphate is one of the major components of the extra-cellular matrix of the vertebrate connective tissues and a high value molecule, widely employed as active principle in the treatment of osteoarthritis. It is usually obtained by extraction from animal tissues, but the risk of virus contaminations, as well as the scarceness of raw material, makes this productive process unsafe and unable to satisfy the growing market demand. In previous studies a new biotechnological process to produce chondroitin from Escherichia coli K4 capsular polysaccharide was investigated and a 1.4 g·L-1 K4 CPS concentration was reached using fed-batch fermentation techniques. In this work, on the trail of these results, we exploited new fermentation strategies to further improve the capsular polysaccharide production. Results The inhibitory effect of acetate on the bacterial cells growth and K4 CPS production was studied in shake flask conditions, while a new approach, that combined the optimization of the feeding profiles, the improvement of aeration conditions and the use of a microfiltration bioreactor, was investigated in three different types of fermentation processes. High polysaccharide concentrations (4.73 ± 0.2 g·L-1, with corresponding average yields (0.13 ± 0.006 gK4 CPS·gcdw-1, were obtained; the increase of K4 CPS titre, compared to batch and fed-batch results, was of 16-fold and 3.3-fold respectively, while average yield was almost 3.5 and 1.4 fold higher. Conclusion The increase of capsular polysaccharide titre confirmed the validity of the proposed fermentation strategy and opened the way to the use of the microfiltration bioreactor for the biotechnological production of chondroitin.

  7. Growth and stress of dourado cultivated in cages at different stocking densities

    Directory of Open Access Journals (Sweden)

    Neiva Braun

    2013-08-01

    Full Text Available The objective of this work was to evaluate the growth and the stress levels of juvenile dourado (Salminus brasiliensis cultivated in cages. Fish stocked at densities of 15 (D15 and 30 (D30 fish per square meter were evaluated in a completely randomized design with three replicates. Fish were fed twice a day with extruded ration (42% crude protein. Density influenced only biomass and daily food intake, and glucose and lactate concentrations increased over time. D15 and D30 did not influence the growth of dourado. However, the increase of glucose and lactate levels over time indicates that cultivation in cages is a stressful condition for this species.

  8. Improved poliovirus D-antigen yields by application of different Vero cell cultivation methods.

    Science.gov (United States)

    Thomassen, Yvonne E; Rubingh, Olaf; Wijffels, René H; van der Pol, Leo A; Bakker, Wilfried A M

    2014-05-19

    Vero cells were grown adherent to microcarriers (Cytodex 1; 3 g L(-1)) using animal component free media in stirred-tank type bioreactors. Different strategies for media refreshment, daily media replacement (semi-batch), continuous media replacement (perfusion) and recirculation of media, were compared with batch cultivation. Cell densities increased using a feed strategy from 1×10(6) cells mL(-1) during batch cultivation to 1.8, 2.7 and 5.0×10(6) cells mL(-1) during semi-batch, perfusion and recirculation, respectively. The effects of these different cell culture strategies on subsequent poliovirus production were investigated. Increased cell densities allowed up to 3 times higher D-antigen levels when compared with that obtained from batch-wise Vero cell culture. However, the cell specific D-antigen production was lower when cells were infected at higher cell densities. This cell density effect is in good agreement with observations for different cell lines and virus types. From the evaluated alternative culture methods, application of a semi-batch mode of operations allowed the highest cell specific D-antigen production. The increased product yields that can easily be reached using these higher cell density cultivation methods, showed the possibility for better use of bioreactor capacity for the manufacturing of polio vaccines to ultimately reduce vaccine cost per dose. Further, the use of animal-component-free cell- and virus culture media shows opportunities for modernization of human viral vaccine manufacturing. Copyright © 2014. Published by Elsevier Ltd.

  9. Development of Cell Analysis Software for Cultivated Corneal Endothelial Cells.

    Science.gov (United States)

    Okumura, Naoki; Ishida, Naoya; Kakutani, Kazuya; Hongo, Akane; Hiwa, Satoru; Hiroyasu, Tomoyuki; Koizumi, Noriko

    2017-11-01

    To develop analysis software for cultured human corneal endothelial cells (HCECs). Software was designed to recognize cell borders and to provide parameters such as cell density, coefficient of variation, and polygonality of cultured HCECs based on phase contrast images. Cultured HCECs with high or low cell density were incubated with Ca-free and Mg-free phosphate-buffered saline for 10 minutes to reveal the cell borders and were then analyzed with software (n = 50). Phase contrast images showed that cell borders were not distinctly outlined, but these borders became more distinctly outlined after phosphate-buffered saline treatment and were recognized by cell analysis software. The cell density value provided by software was similar to that obtained using manual cell counting by an experienced researcher. Morphometric parameters, such as the coefficient of variation and polygonality, were also produced by software, and these values were significantly correlated with cell density (Pearson correlation coefficients -0.62 and 0.63, respectively). The software described here provides morphometric information from phase contrast images, and it enables subjective and noninvasive quality assessment for tissue engineering therapy of the corneal endothelium.

  10. Dynamic cell culture system: a new cell cultivation instrument for biological experiments in space

    Science.gov (United States)

    Gmunder, F. K.; Nordau, C. G.; Tschopp, A.; Huber, B.; Cogoli, A.

    1988-01-01

    The prototype of a miniaturized cell cultivation instrument for animal cell culture experiments aboard Spacelab is presented (Dynamic cell culture system: DCCS). The cell chamber is completely filled and has a working volume of 200 microliters. Medium exchange is achieved with a self-powered osmotic pump (flowrate 1 microliter h-1). The reservoir volume of culture medium is 230 microliters. The system is neither mechanically stirred nor equipped with sensors. Hamster kidney (Hak) cells growing on Cytodex 3 microcarriers were used to test the biological performance of the DCCS. Growth characteristics in the DCCS, as judged by maximal cell density, glucose consumption, lactic acid secretion and pH, were similar to those in cell culture tubes.

  11. [Xenogeneic protein free cultivation of mesenchymal multipotent stromal cells].

    Science.gov (United States)

    Stehlík, D; Pytlík, R; Rychtrmocová, H; Veselá, R; Kopečný, Z; Trč, T

    2011-01-01

    The aim of this study was to compare the standard laboratory method of cultivation of mesenchymal multipotent stromal cells (MSC) and a novel technique of rapid MSC expansion focused on simple clinical use. Bone marrow mononuclear cells of donors were cultured for 14 days by the standard and the new cultivation method. The standard method (STD) was based on an alpha MEM medium supplemented with foetal calf serum (FCS). The new animal protein-free method (CLI) was based on the clinical grade medium CellgroTM, pooled human serum and human recombinant growth factors (EGF, PDGF-BB, M-CSF, FGF-2) supplemented with dexamethasone, insulin and ascorbic acid. The cell product was analyzed by flow cytometry. Furthermore, the cell products of STD and CLI methods were differentiated in vitro, and histochemical and immunohistochemical analyses, electron microscopy and elemental analysis were performed. Some cells were seeded on biodegradable scaffolds, in vivo implanted into immunodeficient mice for 6 weeks and evaluated by histological methods. Yields of the CLI method after 14 days of cultivation were 40-fold higher than those obtained by the STD technique (psystem cultivation in good manufacturing practice (GMP) conditions. It seems to be easily transferable to good clinical practice compared to other protocols and should extend the possibilities of cell therapy and tissue engineering of cartilage and bone. The new method is protected by Czech patent 301 148 and by Europian patent EP 1999250 according to Czech and international laws.

  12. Sorting cells by their density.

    Directory of Open Access Journals (Sweden)

    Nazila Norouzi

    Full Text Available Sorting cells by their type is an important capability in biological research and medical diagnostics. However, most cell sorting techniques rely on labels or tags, which may have limited availability and specificity. Sorting different cell types by their different physical properties is an attractive alternative to labels because all cells intrinsically have these physical properties. But some physical properties, like cell size, vary significantly from cell to cell within a cell type; this makes it difficult to identify and sort cells based on their sizes alone. In this work we continuously sort different cells types by their density, a physical property with much lower cell-to-cell variation within a cell type (and therefore greater potential to discriminate different cell types than other physical properties. We accomplish this using a 3D-printed microfluidic chip containing a horizontal flowing micron-scale density gradient. As cells flow through the chip, Earth's gravity makes each cell move vertically to the point where the cell's density matches the surrounding fluid's density. When the horizontal channel then splits, cells with different densities are routed to different outlets. As a proof of concept, we use our density sorter chip to sort polymer microbeads by their material (polyethylene and polystyrene and blood cells by their type (white blood cells and red blood cells. The chip enriches the fraction of white blood cells in a blood sample from 0.1% (in whole blood to nearly 98% (in the output of the chip, a 1000x enrichment. Any researcher with access to a 3D printer can easily replicate our density sorter chip and use it in their own research using the design files provided as online Supporting Information. Additionally, researchers can simulate the performance of a density sorter chip in their own applications using the Python-based simulation software that accompanies this work. The simplicity, resolution, and throughput of this

  13. Sorting cells by their density.

    Science.gov (United States)

    Norouzi, Nazila; Bhakta, Heran C; Grover, William H

    2017-01-01

    Sorting cells by their type is an important capability in biological research and medical diagnostics. However, most cell sorting techniques rely on labels or tags, which may have limited availability and specificity. Sorting different cell types by their different physical properties is an attractive alternative to labels because all cells intrinsically have these physical properties. But some physical properties, like cell size, vary significantly from cell to cell within a cell type; this makes it difficult to identify and sort cells based on their sizes alone. In this work we continuously sort different cells types by their density, a physical property with much lower cell-to-cell variation within a cell type (and therefore greater potential to discriminate different cell types) than other physical properties. We accomplish this using a 3D-printed microfluidic chip containing a horizontal flowing micron-scale density gradient. As cells flow through the chip, Earth's gravity makes each cell move vertically to the point where the cell's density matches the surrounding fluid's density. When the horizontal channel then splits, cells with different densities are routed to different outlets. As a proof of concept, we use our density sorter chip to sort polymer microbeads by their material (polyethylene and polystyrene) and blood cells by their type (white blood cells and red blood cells). The chip enriches the fraction of white blood cells in a blood sample from 0.1% (in whole blood) to nearly 98% (in the output of the chip), a 1000x enrichment. Any researcher with access to a 3D printer can easily replicate our density sorter chip and use it in their own research using the design files provided as online Supporting Information. Additionally, researchers can simulate the performance of a density sorter chip in their own applications using the Python-based simulation software that accompanies this work. The simplicity, resolution, and throughput of this technique make

  14. LTCC based bioreactors for cell cultivation

    Science.gov (United States)

    Bartsch, H.; Welker, T.; Welker, K.; Witte, H.; Müller, J.

    2016-01-01

    LTCC multilayers offer a wide range of structural options and flexibility of connections not available in standard thin film technology. Therefore they are considered as material base for cell culture reactors. The integration of microfluidic handling systems and features for optical and electrical capturing of indicators for cell culture growth offers the platform for an open system concept. The present paper assesses different approaches for the creation of microfluidic channels in LTCC multilayers. Basic functions required for the fluid management in bioreactors include temperature and flow control. Both features can be realized with integrated heaters and temperature sensors in LTCC multilayers. Technological conditions for the integration of such elements into bioreactors are analysed. The temperature regulation for the system makes use of NTC thermistor sensors which serve as real value input for the control of the heater. It allows the adjustment of the fluid temperature with an accuracy of 0.2 K. The tempered fluid flows through the cell culture chamber. Inside of this chamber a thick film electrode array monitors the impedance as an indicator for the growth process of 3-dimensional cell cultures. At the system output a flow sensor is arranged to monitor the continual flow. For this purpose a calorimetric sensor is implemented, and its crucial design parameters are discussed. Thus, the work presented gives an overview on the current status of LTCC based fluid management for cell culture reactors, which provides a promising base for the automation of cell culture processes.

  15. Surface modification of closed plastic bags for adherent cell cultivation

    Science.gov (United States)

    Lachmann, K.; Dohse, A.; Thomas, M.; Pohl, S.; Meyring, W.; Dittmar, K. E. J.; Lindenmeier, W.; Klages, C.-P.

    2011-07-01

    In modern medicine human mesenchymal stem cells are becoming increasingly important. However, a successful cultivation of this type of cells is only possible under very specific conditions. Of great importance, for instance, are the absence of contaminants such as foreign microbiological organisms, i.e., sterility, and the chemical functionalization of the ground on which the cells are grown. As cultivation of these cells makes high demands, a new procedure for cell cultivation has been developed in which closed plastic bags are used. For adherent cell growth chemical functional groups have to be introduced on the inner surface of the plastic bag. This can be achieved by a new, atmospheric-pressure plasma-based method presented in this paper. The method which was developed jointly by the Fraunhofer IST and the Helmholtz HZI can be implemented in automated equipment as is also shown in this contribution. Plasma process gases used include helium or helium-based gas mixtures (He + N2 + H2) and vapors of suitable film-forming agents or precursors such as APTMS, DACH, and TMOS in helium. The effect of plasma treatment is investigated by FTIR-ATR spectroscopy as well as surface tension determination based on contact angle measurements and XPS. Plasma treatment in nominally pure helium increases the surface tension of the polymer foil due to the presence of oxygen traces in the gas and oxygen diffusing through the gas-permeable foil, respectively, reacting with surface radical centers formed during contact with the discharge. Primary amino groups are obtained on the inner surface by treatment in mixtures with nitrogen and hydrogen albeit their amount is comparably small due to diffusion of oxygen through the gas-permeable bag, interfering with the plasma-amination process. Surface modifications introducing amino groups on the inner surface turned out to be most efficient in the promotion of cell growth.

  16. In vitro cultivation of Schistosoma japonicum-parasites and cells.

    Science.gov (United States)

    Ye, Qing; Dong, Hui-Fen; Grevelding, Christoph G; Hu, Min

    2013-12-01

    Schistosomiasis is a serious parasitic zoonosis caused by blood-dwelling flukes of the genus Schistosoma. Understanding functions of genes and proteins of this parasite is important for uncovering this pathogen's complex biology, which will provide valuable information to design new strategies for schistosomiasis control. Effective applications of molecular tools reported to investigate schistosome gene function, such as inhibitor studies and transgenesis, rely on the developments of in vitro cultivation system of this parasite and cells. Besides the in vitro culture studies dealing with Schistosoma mansoni, there are also numerous excellent studies about the in vitro cultivation of Schistosoma japonicum, which were performed by Chinese researchers and published in Chinese journals. Nearly every stage of the life-cycle of S. japonicum, including miracidia, mother sporocysts, cercariae, schistosomula, and egg-laying adult worms, was employed for developing in vitro cultivation methods, being accompanied by the introduction of several media and supplements that helped to improve culture conditions. It was not only possible to generate mother sporocysts from miracidia in vitro, but also to obtain adult worms from cercariae through in vitro cultivation. The main obstacles to complete the life cycle of S. japonicum in the lab are the transition from mother sporocysts to cercariae, and the production of fertilized and completely developed eggs by adult worms generated in vitro. With regard to cells from S. japonicum, besides established isolation protocols and morphological observations, media optimizations were conducted by using different chemical reagents, biological supplements and physical treatment. Among these, mutagens like N-methyl-N-nitro-N-nitrosoguanidine and the addition of extracellular matrix were found to be able to induce mitogenic activities. Although enzyme activities or the level of silver-stained nucleolar region associated protein in cultured cells

  17. Amino acids in the cultivation of mammalian cells.

    Science.gov (United States)

    Salazar, Andrew; Keusgen, Michael; von Hagen, Jörg

    2016-05-01

    Amino acids are crucial for the cultivation of mammalian cells. This importance of amino acids was realized soon after the development of the first cell lines, and a solution of a mixture of amino acids has been supplied to cultured cells ever since. The importance of amino acids is further pronounced in chemically defined mammalian cell culture media, making the consideration of their biological and chemical properties necessary. Amino acids concentrations have been traditionally adjusted to their cellular consumption rates. However, since changes in the metabolic equilibrium of amino acids can be caused by changes in extracellular concentrations, metabolomics in conjunction with flux balance analysis is being used in the development of culture media. The study of amino acid transporters is also gaining importance since they control the intracellular concentrations of these molecules and are influenced by conditions in cell culture media. A better understanding of the solubility, stability, dissolution kinetics, and interactions of these molecules is needed for an exploitation of these properties in the development of dry powdered chemically defined media for mammalian cells. Due to the complexity of these mixtures however, this has proven to be challenging. Studying amino acids in mammalian cell culture media will help provide a better understanding of how mammalian cells in culture interact with their environment. It would also provide insight into the chemical behavior of these molecules in solutions of complex mixtures, which is important in the understanding of the contribution of individual amino acids to protein structure.

  18. Reimplantation of cultivated human bone cells from the posterior maxilla for sinus floor augmentation

    DEFF Research Database (Denmark)

    Hermund, Niels Ulrich; Stavropoulos, Andreas; Donatsky, Ole

    2012-01-01

    OBJECTIVES: The aim of the present randomized clinical study was to evaluate histologically whether the addition of cultivated, autogenous bone cells to a composite graft of deproteinized bovine bone mineral (DBBM) and autogenous bone (AB) for sinus floor augmentation (SFA) enhance bone formation...... compared with what achieved after SFA with DBBM + AB alone. MATERIAL AND METHODS: Twenty patients with remaining posterior maxillary alveolar crest height of less than 3 mm received SFA after randomization either with an DBBM and AB composite in a 1 : 1 ratio or with DBBM + AB supplemented with autogenous...... the tuberosity area. Bone density at the augmented sinus and the tuberosity area and the height of augmentation were estimated on non-decalcified histological sections prepared from the biopsies. A relative bone density index (RBD) was also calculated by dividing bone density at the augmented sinus with bone...

  19. High-density cultivation of Lactobacillus plantarum NCU116 in an ...

    African Journals Online (AJOL)

    In this study, we reported a simple fedbatch process for Lactobacillus plantarum NCU116 with high cell density. It was found that the optimum initial glucose concentration for this strain was 5% (w/v). To reduce the effect of acid and starvation, the exponential fed-batch culture and ammonium fed-batch system were ...

  20. Stem Cells from Human Exfoliated Deciduous Teeth – Isolation, Long Term Cultivation and Phenotypical Analysis

    Directory of Open Access Journals (Sweden)

    Jakub Suchánek

    2010-01-01

    Full Text Available Aims: Our aims were to isolate stem cells from human exfoliated deciduous teeth (SHED, to cultivate them in vitro and to investigate their basic biological properties, phenotype and to compare our findings with dental pulp stem cells (DPSC isolated from permanent teeth. Methods: Dental pulp was gently evacuated from exfoliated teeth. After enzymatic dissociation of dental pulp, SHED were cultivated in modified cultivation media for mesenchymal adult progenitor cells containing 2 % FCS and supplemented with growth factors and insulin, transferrin, sodium (ITS supplement. Cell viability and other biological properties were examined using a Vi-Cell analyzer and a Z2-Counter. DNA analyses and phenotyping were performed with flow cytometry. Results: We were able to cultivate SHED over 45 population doublings. Our results showed that SHED cultivated under same conditions as DPSC had longer average population doubling time (41.3 hrs for SHED vs. 24.5 hrs for DPSC. Phenotypic comparison of cultivated SHED to that of cultivated DPSC showed differential expression CD29, CD44, CD71, CD117, CD166. During long-term cultivation, SHED did not showed any signs of degeneration or spontaneous differentiation. Conclusions: We isolated stem cells from exfoliated teeth. In comparison to DPSC, SHED proliferation rate was about 50% slower, and SHED showed slightly different phenotype. These cells may be extremely useful for stem cell tissue banking, further stem cell research and future therapeutic applications.

  1. Cultivation and Biological Characterization of Chicken Primordial Germ Cells

    Directory of Open Access Journals (Sweden)

    Meng Ji

    2016-01-01

    Full Text Available The purpose of this work was to investigate the isolation, culture process of chicken gonadal primordial germ cells (PGCs and study their biological characterization. PGCs were harvested from 5.5-day-old chicken embryonic genital ridges and explanted onto chicken embryonic fibroblasts (CEFs. The results showed that the primary cultivation of chicken PGCs on their own gonadal stroma cells were better than CEFs at first two days for reproduction. The conditioned media supported the growth and colony formation of PGCs for a prolonged time in vitro and maintained a normal diploid karyotype, which were positively stained by alkaline phosphatase (AKP, periodic acid Schiff (PAS and reacted with anti-SSEA-1, SSEA-3, Oct4, Blimp1 and Sox2. Real-time PCR showed that they expressed the stage specific genes CVH, Blimp1 and Dazl, the stem cell specific genes Sox2, Pouv and Nanog. They also formed the embryoid bodies (EBs. These results suggested that the chicken PGCs cultured in vitro not only had strong self-renewal ability, but also had the potential capability of multi-lineage differentiation.

  2. Effects of seedling age and cultivation density on agronomic characteristics and grain yield of mechanically transplanted rice.

    Science.gov (United States)

    Liu, Qihua; Zhou, Xuebiao; Li, Jingling; Xin, Caiyun

    2017-10-26

    Delayed transplantation frequently occurs in mechanically transplanted rice in China, leading to a significant reduction in grain yield. Thus, determining how to compensate grain yield loss is crucial for improving rice cultivation technology. A field experiment was conducted to investigate the effects of cultivation density and seedling age on agronomic traits and grain yield of mechanically transplanted rice. With increasing seedling age, rice tiller number, pre-anthesis dry matter accumulation, remobilization efficiency and contribution to grain yield, as well as post-anthesis photosynthesis amount decreased, causing reductions in the number of effective panicles, the total number of grains per panicle, the sink capacity per tiller, and grain yield. In rice transplanted at 30- and 35-day seedling ages, increasing cultivation density significantly enhanced the number of effective panicles and grain yield. Additionally, there existed strong, positive correlations between sink capacity per tiller and pre-anthesis dry matter remobilization efficiency and pre-anthesis dry matter contribution to grain yield. We conclude that in addition to cultivation density, enhancing the amount of pre-anthesis dry matter and the remobilization efficiency could be feasible for mitigating grain yield loss caused by delayed transplantation.

  3. Effects of plant densities on yield, yield components and some morphological characters of two cultivators of oilseed rape (Brassica napus L.)

    DEFF Research Database (Denmark)

    Al-Barzinjy, M.; Stölen, O.; Christiansen, Jørgen Lindskrog

    2003-01-01

    Effects of Plant Densities on Yield, Yield Components and some Morphological Characters of two Cultivators of Oilseed Rape (Brassica napus L.)......Effects of Plant Densities on Yield, Yield Components and some Morphological Characters of two Cultivators of Oilseed Rape (Brassica napus L.)...

  4. [The cultivation of bone marrow cells and cell lines on polymeric films].

    Science.gov (United States)

    Dolgikh, M S; Livak, D N; Krasheninnikov, M E; Onishchenko, N A

    2011-01-01

    The cultivation of multipotent mesenchymal stromal bone marrow cells and cells of A-431, MDCK, Vero, 3T3 and Hep-G2 was performed on polymeric films (PVA) with different hydrophobic fatty acid residues. The cells of different types grew on these films with different intensity, but in the most cases comparable with the cultivation control on usual plastic. The examined films were nontoxic to cells and sufficiently adhesive. They did not changed pH of cultural media, were optically transparent under microscope and comfortable in the experimental work. These films can be used as a model for the artificial organ construction. The covalent binding of different fatty acids to PVA shows possibility of the adaptable changes of films properties (hydrophobity and adhesiveness), and therefore possibility of the creation of optimal conditions for different cell types attachement and growth.

  5. Bacterial density and survey of cultivable heterotrophs in the surface water of a freshwater marsh habitat of Anopheles quadrimaculatus larvae (Diptera: Culicidae).

    Science.gov (United States)

    Smith, T W; Walker, E D; Kaufman, M G

    1998-03-01

    We examined surface water samples collected in September and October 1994 from a freshwater marsh habitat containing larval Anopheles quadrimaculatus mosquitoes. Bacterial densities in direct microscopic counts ranged from 9.7 x 10(5) to 1.3 x 10(7) cells/ml. Densities of cultivable bacteria on trypticase soy agar medium ranged from 1.0 to 1.5 x 10(5) cells/ml. The majority of 888 isolates were gram-positive rods (41%) followed by gram-negative rods (28%). Analysis of the cellular fatty acid profiles of 824 isolates using gas chromatography and Microbial Identification Systems TSBA (Rev. 3.60) library software grouped the bacteria into Bacillus spp. (35%), other gram-positive bacteria (16%), pseudomonads (15%), other gram-negative bacteria including mainly Enterobacteriaceae and Vibrionaceae (21%), and profiles not recognized (13%). Among 33 genera within these groups, the most common were Bacillus, Pseudomonas, Aeromonas, and Arthrobacter.

  6. Shaping of Natural Killer Cell Antitumor Activity by Ex Vivo Cultivation

    Science.gov (United States)

    Granzin, Markus; Wagner, Juliane; Köhl, Ulrike; Cerwenka, Adelheid; Huppert, Volker; Ullrich, Evelyn

    2017-01-01

    Natural killer (NK) cells are a promising tool for the use in adoptive immunotherapy, since they efficiently recognize and kill tumor cells. In this context, ex vivo cultivation is an attractive option to increase NK cells in numbers and to improve their antitumor potential prior to clinical applications. Consequently, various strategies to generate NK cells for adoptive immunotherapy have been developed. Here, we give an overview of different NK cell cultivation approaches and their impact on shaping the NK cell antitumor activity. So far, the cytokines interleukin (IL)-2, IL-12, IL-15, IL-18, and IL-21 are used to culture and expand NK cells. The selection of the respective cytokine combination is an important factor that directly affects NK cell maturation, proliferation, survival, distribution of NK cell subpopulations, activation, and function in terms of cytokine production and cytotoxic potential. Importantly, cytokines can upregulate the expression of certain activating receptors on NK cells, thereby increasing their responsiveness against tumor cells that express the corresponding ligands. Apart from using cytokines, cocultivation with autologous accessory non-NK cells or addition of growth-inactivated feeder cells are approaches for NK cell cultivation with pronounced effects on NK cell activation and expansion. Furthermore, ex vivo cultivation was reported to prime NK cells for the killing of tumor cells that were previously resistant to NK cell attack. In general, NK cells become frequently dysfunctional in cancer patients, for instance, by downregulation of NK cell activating receptors, disabling them in their antitumor response. In such scenario, ex vivo cultivation can be helpful to arm NK cells with enhanced antitumor properties to overcome immunosuppression. In this review, we summarize the current knowledge on NK cell modulation by different ex vivo cultivation strategies focused on increasing NK cytotoxicity for clinical application in malignant

  7. Increasing Vero viable cell densities for yellow fever virus production in stirred-tank bioreactors using serum-free medium.

    Science.gov (United States)

    Mattos, Diogo A; Silva, Marlon V; Gaspar, Luciane P; Castilho, Leda R

    2015-08-20

    In this work, changes in Vero cell cultivation methods have been employed in order to improve cell growth conditions to obtain higher viable cell densities and to increase viral titers. The propagation of the 17DD yellow fever virus (YFV) in Vero cells grown on Cytodex I microcarriers was evaluated in 3-L bioreactor vessels. Prior to the current changes, Vero cells were repeatedly displaying insufficient microcarrier colonization. A modified cultivation process with four changes has resulted in higher cell densities and higher virus titers than previously observed for 17DD YFV. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Formation of industrial mixed culture biofilm in chlorophenol cultivated medium of microbial fuel cell

    Science.gov (United States)

    Hassan, Huzairy; Jin, Bo; Dai, Sheng; Ngau, Cornelius

    2016-11-01

    The formation of microbial biofilm while maintaining the electricity output is a challenging topic in microbial fuel cell (MFC) studies. This MFC critical factor becomes more significant when handling with industrial wastewater which normally contains refractory and toxic compounds. This study explores the formation of industrial mixed culture biofilm in chlorophenol cultivated medium through observing and characterizing microscopically its establishment on MFC anode surface. The mixed culture was found to develop its biofilm on the anode surface in the chlorophenol environment and established its maturity and dispersal stages with concurrent electricity generation and phenolic degradation. The mixed culture biofilm engaged the electron transfer roles in MFC by generating current density of 1.4 mA/m2 and removing 53 % of 2,4-dichlorophenol. The results support further research especially on hazardous wastewater treatment using a benign and sustainable method.

  9. Analysing the Influence of the Spontaneous Aneuploidy Frequency on the Cell Population System Cultivation

    Directory of Open Access Journals (Sweden)

    G. A. Nefedov

    2015-01-01

    Full Text Available The paper provides a qualitative analysis of M.S. Vinogradova's nonlinear model for dynamics of the cell population system. This system describes the stem cells cultivation in vitro under resource constraints. The system consists of two populations, namely: population of normal cells and population of abnormal cells. Resource constraints are considered as linear dependences of mitosis parameters on the normalized densities of each population.One of the key parameters that effects on the realization of the system evolution scenarios is a parameter that determines a share of the normal cells, which pass, when dividing, into population of the abnormal cells. The paper analyses both the existence conditions of the rest points and the changes of the evolution scenarios of population system with changing abovementioned parameter and other system parameters held fixed. It is shown that there is a saddle-node bifurcation in the system; the bifurcation value of the parameter is found. The paper shows the interval of parameter values in which the favorable scenarios of population system evolution are implemented. It also presents results of mathematical modeling.

  10. Bone Proteomics experiment (BOP): the first proteomics analysis of mammalian cells cultivated in weightlessness conditions

    Science.gov (United States)

    Costessi, A.; Vascotto, C.; Pines, A.; Romanello, M.; Schonenborg, R.; Schiller, P.; Moro, L.; Tell, G.

    Bone mass loss is a major consequence of extended periods of weightlessness Many studies have been performed on astronauts and animal models establishing that a decrease of the maturation process and of the bone synthesising activity of osteoblast cells play a key role in microgravity-dependent bone mass loss Several experiments on single cells and tissues showed that weightlessness can also influence cells cultivated in vitro Many molecular mechanisms are affected among which the cytoskeleton and intracellular signal transduction cascades However the underlying mechanisms of these changes and their molecular consequences are far from being fully understood and the cellular gravisensing machinery is still unknown In contrast to weightlessness dynamic mechanical loading increases bone density and strength and promotes osteoblast proliferation differentiation and matrix production by acting at the gene expression level However the molecular mechanisms by which mechanical forces are converted into biochemical signalling in bone are also poorly understood A growing body of evidence points to extracellular nucleotides i e ATP and UTP as soluble factors that are released by several cell types in response to mechanical stimulation and that eventually trigger an intracellular signal We have recently demonstrated in the HOBIT osteoblast cell line that ATP and UTP treatments can activate two fundamental transcription factors that promote osteoblast differentiation and physiology Runx2 and Egr-1 as well as their target genes galectin-3 and

  11. Shape-controlled high cell-density microcapsules by electrodeposition.

    Science.gov (United States)

    Liu, Zeyang; Takeuchi, Masaru; Nakajima, Masahiro; Hasegawa, Yasuhisa; Huang, Qiang; Fukuda, Toshio

    2016-06-01

    Cell encapsulation within alginate-poly-l-lysine (PLL) microcapsules has been developed to provide a miniaturized three-dimensional (3D) microenvironment with an aqueous core while promoting development of encapsulated cells into high cell-density structures. In this paper, a novel method for fabricating shape-controlled alginate-PLL microcapsules to construct 3D cell structures based on electrodeposition method is provided. Two-dimensional Ca-alginate cell-laden gel membranes were electrodeposited onto a micro-patterned electrode and further detached from the electrode. The PLL was coated onto the gel structures to form alginate-PLL complex as an outer shell and sodium citric solution was utilized to melt the internal alginate to achieve miniaturized 3D microcapsules (sphere, cuboid, and rod shape). By this proposed method, rat liver cells (RLC-18) formed multi-cellular aggregates with high cell-density after cultivation for 2weeks. The use of alginate-poly-l-lysine (PLL) microcapsules has shown great potential in fabricating 3D cell structures with high cell density. Despite their success related to their ability to provide a miniaturized microenvironment with an aqueous core, alginate-PLL microcapsules has drawback such as a limited shape-control ability. Because of the mechanism of Ca-induced alginate gel formation, it is still difficult to precisely control the gelation process to produce alginate-PLL microcapsules with specific shape. The present study provides an electrodeposition-based method to generate shape-controlled microcapsules for 3D cell structures. Sphere, cuboid, and rod shaped microcapsules of RLC-18 cells were produced for long-term culture to obtain desired morphologies of cell aggregates. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  12. Photo-cross-linking of amniotic membranes for limbal epithelial cell cultivation

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Jui-Yang, E-mail: jylai@mail.cgu.edu.tw

    2014-12-01

    In the present study, we developed photo-cross-linked amniotic membrane (AM) as a limbal stem cell niche. After ultraviolet (UV) irradiation for varying time periods, the biological tissues were studied by determinations of cross-linking structure, degradability, and nutrient permeation ability. Our results showed that the number of cross-links per unit mass of AM significantly increased with increasing illumination time from 5 to 50 min. However, the cross-link formation was inhibited by longer irradiation time (i.e., 150 min), probably due to the scission of tissue collagen chains through irradiation. The biological stability and matrix permeability of photo-cross-linked AM materials strongly depended on their cross-linking densities affected by the UV irradiation. In vitro biocompatibility studies including cell viability and pro-inflammatory gene expression analyses demonstrated that, irrespective of the irradiation time employed, the physically cross-linked biological tissues exhibited negligible cytotoxicity and similar interleukin-6 (IL-6) mRNA levels. The data clearly indicate that these AM matrices do not cause potential harm to the corneal epithelial cells. After the growth of limbal epithelial cells (LECs) on AM substrates, Western blot analyses were conducted to examine the expression of ABCG2. It was found that the ability of UV-irradiated AM to maintain the undifferentiated precursor cell phenotype was significantly enhanced with increasing extent of photo-cross-linking. In summary, the UV irradiation time may have a profound influence on the fabrication of photo-cross-linked AM matrices for LEC cultivation. - Highlights: • We report the development of photo-cross-linked AM as a limbal stem cell niche. • Cross-linked structure of tissue materials was controlled by UV irradiation time. • Biostability and matrix permeability of AM depended on cross-linking density. • All the studied photo-cross-linked AM showed good in vitro biocompatibility.

  13. Behaviour of human umbilical vein endothelial cells (HUVEC) cultivated in microfluidic channels

    NARCIS (Netherlands)

    Mulder, Patty P. M. F. A.; Molema, Grietje; Koster, Sander; van der Linden, Heiko J.; Verpoorte, Elisabeth

    2006-01-01

    Our long-term goal is to develop advanced tools for cell studies and analysis based on microfluidic systems. In this paper, we report on endothelial cell cultivation in microchannels and 96-well tissue plates, and compare cell phenotype and cellular status in the two enviroments. This was done under

  14. High-density polymer microarrays: identifying synthetic polymers that control human embryonic stem cell growth.

    Science.gov (United States)

    Hansen, Anne; Mjoseng, Heidi K; Zhang, Rong; Kalloudis, Michail; Koutsos, Vasileios; de Sousa, Paul A; Bradley, Mark

    2014-06-01

    The fabrication of high-density polymer microarray is described, allowing the simultaneous and efficient evaluation of more than 7000 different polymers in a single-cellular-based screen. These high-density polymer arrays are applied in the search for synthetic substrates for hESCs culture. Up-scaling of the identified hit polymers enables long-term cellular cultivation and promoted successful stem-cell maintenance. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Featured Article: Isolation, characterization, and cultivation of human hepatocytes and non-parenchymal liver cells

    Science.gov (United States)

    Pfeiffer, Elisa; Kegel, Victoria; Zeilinger, Katrin; Hengstler, Jan G; Nüssler, Andreas K; Seehofer, Daniel

    2015-01-01

    Primary human hepatocytes (PHH) are considered to be the gold standard for in vitro testing of xenobiotic metabolism and hepatotoxicity. However, PHH cultivation in 2D mono-cultures leads to dedifferentiation and a loss of function. It is well known that hepatic non-parenchymal cells (NPC), such as Kupffer cells (KC), liver endothelial cells (LEC), and hepatic stellate cells (HSC), play a central role in the maintenance of PHH functions. The aims of the present study were to establish a protocol for the simultaneous isolation of human PHH and NPC from the same tissue specimen and to test their suitability for in vitro co-culture. Human PHH and NPC were isolated from tissue obtained by partial liver resection by a two-step EDTA/collagenase perfusion technique. The obtained cell fractions were purified by Percoll density gradient centrifugation. KC, LEC, and HSC contained in the NPC fraction were separated using specific adherence properties and magnetic activated cell sorting (MACS®). Identified NPC revealed a yield of 1.9 × 106 KC, 2.7 × 105 LEC and 4.7 × 105 HSC per gram liver tissue, showing viabilities >90%. Characterization of these NPC showed that all populations went through an activation process, which influenced the cell fate. The activation of KC strongly depended on the tissue quality and donor anamnesis. KC became activated in culture in association with a loss of viability within 4–5 days. LEC lost specific features during culture, while HSC went through a transformation process into myofibroblasts. The testing of different culture conditions for HSC demonstrated that they can attenuate, but not prevent dedifferentiation in vitro. In conclusion, the method described allows the isolation and separation of PHH and NPC in high quality and quantity from the same donor. PMID:25394621

  16. Production Process for Stem Cell Based Therapeutic Implants: Expansion of the Production Cell Line and Cultivation of Encapsulated Cells

    Science.gov (United States)

    Weber, C.; Pohl, S.; Poertner, R.; Pino-Grace, Pablo; Freimark, D.; Wallrapp, C.; Geigle, P.; Czermak, P.

    Cell based therapy promises the treatment of many diseases like diabetes mellitus, Parkinson disease or stroke. Microencapsulation of the cells protects them against host-vs-graft reactions and thus enables the usage of allogenic cell lines for the manufacturing of cell therapeutic implants. The production process of such implants consists mainly of the three steps expansion of the cells, encapsulation of the cells, and cultivation of the encapsulated cells in order to increase their vitality and thus quality. This chapter deals with the development of fixed-bed bioreactor-based cultivation procedures used in the first and third step of production. The bioreactor system for the expansion of the stem cell line (hMSC-TERT) is based on non-porous glass spheres, which support cell growth and harvesting with high yield and vitality. The cultivation process for the spherical cell based implants leads to an increase of vitality and additionally enables the application of a medium-based differentiation protocol.

  17. In vivo osteoinductive effect and in vitro isolation and cultivation bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Redzić, Amira; Smajilagić, Amer; Aljicević, Mufida; Berberović, Ljubomir

    2010-12-01

    Bone marrow contains cell type termed mesenchymal stem cells (MSC), first recognized in bone marrow by a German pathologist, Julius Cohnheim in 1867. That MSCs have potential to differentiate in vitro in to the various cells lines as osteoblast, chondroblast, myoblast and adipoblast cells lines. Aims of our study were to show in vivo capacity of bone marrow MSC to produce bone in surgically created non critical size mandible defects New Zeland Rabbits, and then in second part of study to isolate in vitro MSC from bone marrow, as potential cell transplantation model in bone regeneration. In vivo study showed new bone detected on 3D CT reconstruction day 30, on all 3 animals non critical size defects, treated with bone marrow MSC exposed to the human Bone Morphogenetic Protein 7 (rhBMP-7). Average values of bone mineral density (BMD), was 530 mg/cm3, on MSC treated animals, and 553 mg/cm3 on control group of 3 animals where non critical size defects were treated with iliac crest autologue bone graft. Activity of the Alkaline Phosphatase enzyme were measurement on 0.5, 14, 21, 30 day and increased activity were detected day 14 on animals treated with bone marrow MSCs compared with day 30 on iliac crest treated animals. That results indicates strong osteoinduction activity of the experimental bone marrow MSCs models exposed to the rhBMP-7 factor Comparing ALP activity, that model showed superiorly results than control group. That result initiates us in opinion that MSCs alone should be alternative for the autolologue bone transplantation and in vitro study we isolated singles MSCs from the bone marrow of rat's tibia and femora and cultivated according to the method of Maniatopoulos et all. The small initial colonies of fibroblast like cells were photo-documented after 2 days of primary culture. Such isolated and cultivated MSCs in future studies will be exposed to the growth factors to differentiate in osteoblast and indicate their clinically potential as alternative

  18. Algae-facilitated chemical phosphorus removal during high-density Chlorella emersonii cultivation in a membrane bioreactor.

    Science.gov (United States)

    Xu, Meng; Bernards, Matthew; Hu, Zhiqiang

    2014-02-01

    An algae-based membrane bioreactor (A-MBR) was evaluated for high-density algae cultivation and phosphorus (P) removal. The A-MBR was seeded with Chlorella emersonii and operated at a hydraulic retention time of 1day with minimal biomass wastage for about 150days. The algae concentration increased from initially 385mg/L (or 315mg biomass COD/L) to a final of 4840mg/L (or 1664mg COD/L), yielding an average solids (algae biomass+minerals) production rate of 32.5gm(-3)d(-1) or 6.2gm(-2)d(-1). The A-MBR was able to remove 66±9% of the total P from the water while the algal biomass had an average of 7.5±0.2% extracellular P and 0.4% of intracellular P. The results suggest that algae-induced phosphate precipitation by algae is key to P removal and high-density algae cultivation produces P-rich algal biomass with excellent settling properties. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. A novel horizontal photobioreactor for high-density cultivation of microalgae.

    Science.gov (United States)

    Dogaris, Ioannis; Welch, Michael; Meiser, Andreas; Walmsley, Lawrence; Philippidis, George

    2015-12-01

    Microalgae are a promising source of biofuels and bioproducts, as they consume CO2 to grow, multiply quickly, and can be cultivated in wastewater and on marginal land. Development of low-cost and high-efficiency microalgal cultivation systems is important to the cost-competitiveness of algae. A floating horizontal photobioreactor (HBR) was developed that is inexpensive and scalable, as it is manufactured from inexpensive plastic film and is modular. Its performance was successfully tested using the marine microalgae Nannochloris atomus Butcher CCAP 251/4A in a 65-L prototype unit. High biomass concentration of 4.0 g L(-1) and productivity of 12.9 g m(-2)d(-1) was achieved indoors under artificial illumination of 31.3 klux (435 μmol m(-2)s(-1)). Outdoors, during semi-continuous operation in Florida, the HBR achieved over the course of 165 days a maximum biomass concentration of 4.3 g L(-1) and an average biomass productivity of 18.2 g m(-2)d(-1) without any contamination issues. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Susceptibility of various cell lines to Neospora caninum tachyzoites cultivation

    Directory of Open Access Journals (Sweden)

    Khordadmehr, M.,

    2014-05-01

    Full Text Available Neospora caninum is a coccidian protozoan parasite which is a major cause of bovine abortions and neonatal mortality in cattle, sheep, goat and horse. Occasionally, cultured cells are used for isolation and multiplication of the agent in vitro with several purposes. In this study the tachyzoite yields of N. caninum were compared in various cell cultures as the host cell lines. Among the cell cultures tested, two presented good susceptibility to the agent: cell lines Vero and MA-104. SW742 and TLI (in vitro suspension culture of lymphoid cells infected with Theileria lestoquardi showed moderate sensitivity. No viable tachyzoite were detected in the culture of MDCK and McCoy cell lines. These results demonstrate that MA-104 and SW742 cells present adequate susceptibility to N. caninum compared to Vero cells, which have been largely used to multiply the parasite in vitro. Moreover, these have easy manipulation, fast multiplication and relatively low nutritional requirements. In addition, the result of this study showed that TLI cell line as a suspension cell culture is susceptible to Nc-1 tachyzoites infection and could be used as an alternative host cell line for tachyzoites culture in vitro studies.

  1. Five willow varieties cultivated across diverse field environments reveal stem density variation associated with high tension wood abundance

    Directory of Open Access Journals (Sweden)

    Nicolas eBerthod

    2015-10-01

    Full Text Available Sustainable and inexpensive production of biomass is necessary to make biofuel production feasible, but represents a challenge. Five short rotation coppice (SRC willow cultivars, selected for high biomass yield, were cultivated on sites at four diverse regions of Quebec to determine their bioenergy potential in contrasting environments. Wood composition and anatomical traits were characterized. Tree height and stem diameter were measured to evaluate growth performance of the cultivars according to the diverse pedoclimatic conditions. Each cultivar showed very specific responses to its environment. While no significant variation in lignin content was observed between sites, there was variation between cultivars. Surprisingly, the pattern of substantial genotype variability in stem density was maintained across all sites. However, wood anatomy did differ between sites in a cultivar (producing high and low density wood, suggesting a probable response to an abiotic stress. Furthermore, twice as many cellulose-rich G-fibers, comprising over 50 % of secondary xylem, were also found in the high density wood, a finding with potential to bring higher value to the lignocellulosic bioethanol industry

  2. Five willow varieties cultivated across diverse field environments reveal stem density variation associated with high tension wood abundance.

    Science.gov (United States)

    Berthod, Nicolas; Brereton, Nicholas J B; Pitre, Frédéric E; Labrecque, Michel

    2015-01-01

    Sustainable and inexpensive production of biomass is necessary to make biofuel production feasible, but represents a challenge. Five short rotation coppice willow cultivars, selected for high biomass yield, were cultivated on sites at four diverse regions of Quebec in contrasting environments. Wood composition and anatomical traits were characterized. Tree height and stem diameter were measured to evaluate growth performance of the cultivars according to the diverse pedoclimatic conditions. Each cultivar showed very specific responses to its environment. While no significant variation in lignin content was observed between sites, there was variation between cultivars. Surprisingly, the pattern of substantial genotype variability in stem density was maintained across all sites. However, wood anatomy did differ between sites in a cultivar (producing high and low density wood), suggesting a probable response to an abiotic stress. Furthermore, twice as many cellulose-rich G-fibers, comprising over 50% of secondary xylem, were also found in the high density wood, a finding with potential to bring higher value to the lignocellulosic bioethanol industry.

  3. Purification and Cultivation of Human Pituitary Growth Hormones Secreting Cells

    Science.gov (United States)

    Hymer, W. C.; Todd, P.; Grindeland, R.; Lanham, W.; Morrison, D.

    1985-01-01

    The rat and human pituitary gland contains a mixture of hormone producing cell types. The separation of cells which make growth hormone (GH) is attempted for the purpose of understanding how the hormone molecule is made within the pituitary cell; what form(s) it takes within the cell; and what form(s) GH assumes as it leaves the cell. Since GH has a number of biological targets (e.g., muscle, liver, bone), the assessment of the activities of the intracellular/extracellular GH by new and sensitive bioassays. GH cells contained in the mixture was separated by free flow electrophoresis. These experiments show that GH cells have different electrophoretic mobilities. This is relevant to NASA since a lack of GH could be a prime causative factor in muscle atrophy. Further, GH has recently been implicated in the etiology of motion sickness in space. Continous flow electrophoresis experiment on STS-8 showed that GH cells could be partially separated in microgravity. However, definitive cell culture studies could not be done due to insufficient cell recoveries.

  4. Biomek Cell Workstation: A Flexible System for Automated 3D Cell Cultivation.

    Science.gov (United States)

    Lehmann, R; Gallert, C; Roddelkopf, T; Junginger, S; Thurow, K

    2016-08-01

    The shift from 2D cultures to 3D cultures enables improvement in cell culture research due to better mimicking of in vivo cell behavior and environmental conditions. Different cell lines and applications require altered 3D constructs. The automation of the manufacturing and screening processes can advance the charge stability, quality, repeatability, and precision. In this study we integrated the automated production of three 3D cell constructs (alginate beads, spheroid cultures, pellet cultures) using the Biomek Cell Workstation and compared them with the traditional manual methods and their consequent bioscreening processes (proliferation, toxicity; days 14 and 35) using a high-throughput screening system. Moreover, the possible influence of antibiotics (penicillin/streptomycin) on the production and screening processes was investigated. The cytotoxicity of automatically produced 3D cell cultures (with and without antibiotics) was mainly decreased. The proliferation showed mainly similar or increased results for the automatically produced 3D constructs. We concluded that the traditional manual methods can be replaced by the automated processes. Furthermore, the formation, cultivation, and screenings can be performed without antibiotics to prevent possible effects. © 2015 Society for Laboratory Automation and Screening.

  5. Growth and metabolism of mesenchymal stem cells cultivated on microcarriers

    NARCIS (Netherlands)

    Schop, Deborah

    2010-01-01

    Mesenchymal stem cells, MSCs, are a great potential source for clinical applications in the field of tissue regeneration. Although MSCs can be isolated from several tissues of the human body, e.g. the bone marrow, the tissues does not contain clinically relevant amounts of MSCs for cell therapeutic

  6. Best practices in heterotrophic high-cell-density microalgal processes: achievements, potential and possible limitations.

    Science.gov (United States)

    Bumbak, Fabian; Cook, Stella; Zachleder, Vilém; Hauser, Silas; Kovar, Karin

    2011-07-01

    Microalgae of numerous heterotrophic genera (obligate or facultative) exhibit considerable metabolic versatility and flexibility but are currently underexploited in the biotechnological manufacturing of known plant-derived compounds, novel high-value biomolecules or enriched biomass. Highly efficient production of microalgal biomass without the need for light is now feasible in inexpensive, well-defined mineral medium, typically supplemented with glucose. Cell densities of more than 100 g l(-1) cell dry weight have been achieved with Chlorella, Crypthecodinium and Galdieria species while controlling the addition of organic sources of carbon and energy in fedbatch mode. The ability of microalgae to adapt their metabolism to varying culture conditions provides opportunities to modify, control and thereby maximise the formation of targeted compounds with non-recombinant microalgae. This review outlines the critical aspects of cultivation technology and current best practices in the heterotrophic high-cell-density cultivation of microalgae. The primary topics include (1) the characteristics of microalgae that make them suitable for heterotrophic cultivation, (2) the appropriate chemical composition of mineral growth media, (3) the different strategies for fedbatch cultivations and (4) the principles behind the customisation of biomass composition. The review confirms that, although fundamental knowledge is now available, the development of efficient, economically feasible large-scale bioprocesses remains an obstacle to the commercialisation of this promising technology.

  7. Human Dental Pulp Stem Cells – Isolation and Long Term Cultivation

    Directory of Open Access Journals (Sweden)

    Jakub Suchánek

    2007-01-01

    Full Text Available Human adult mesenchymal stem cells (MSCs are rare elements living in various organs (e.g. bone marrow, skeletal muscle, with capability to differentiate in various cell types (e.g. chondrocytes, adipocytes and osteoblasts. In the year 2000, Gronthos and co-workers isolated stem cells from the human dental pulp (DPSCs. Later on, stem cells from exfoliated tooth were also obtained. The aims of our study were to establish protocol of DPSCs isolation and to cultivate DPSCs either from adult or exfoliated tooth, and to compare these cells with mesenchymal progenitor cell (MPCs cultures. MPCs were isolated from the human bone marrow of proximal femur. DPSCs were isolated from deciduous and permanent teeth. Both cell types were cultivated under the same conditions in the media with 2 % of FCS supplemented with PDGF and EGF growth factors. We have cultivated undifferentiated DPSCs for long time, over 60 population doublings in cultivation media designed for bone marrow MPCs. After reaching Hayflick’s limit, they still have normal karyotype. Initial doubling time of our cultures was from 12 to 50 hours for first 40 population doublings, after reaching 50 population doublings, doubling time had increased to 60–90 hours. Regression analysis of uncumulated population doublings proved tight dependence of population doublings on passage number and slow decrease of proliferation potential. In comparison with bone marrow MPCs, DPSCs share similar biological characteristics and stem cell properties. The results of our experiments proved that the DPSCs and MPCs are highly proliferative, clonogenic cells that can be expanded beyond Hayflick’s limit and remain cytogenetically stable. Moreover we have probably isolated two different populations of DPSCs. These DPSCs lines differed one from another in morphology. Because of their high proliferative and differentiation potential, DPSCs can become more attractive, easily accessible source of adult stem cells for

  8. Amaranthus cruentus L. is suitable for cultivation in Central Italy: field evaluation and response to plant densities

    Directory of Open Access Journals (Sweden)

    Paolo Casini

    2014-12-01

    Full Text Available The aim of this study was to determine the possibility of amaranth cultivation in Central Italy and to determine the optimum plant density. Field trials were carried out in 2011 and 2012 under non-irrigated conditions in Tuscany (43° 18’ N, 11° 47’ E. Twelve accessions of two amaranth species (Amaranthus cruentus L. and A. hypochondriacus L. were utilised. Genotypes were evaluated over a two-year period using a RCB design with three replicates. The effects of plant density were investigated in 2012. A with a split-plot design was used, where the A. cruentus accessions (AMES 5148, PI 511719 and PI 643045 constituted the main plots. Plant densities (7.5, 15, 30 and 60 plants m–2 constituted the subplots. Plants were transplanted at the 3-4 true leaf stage. Morphological traits were determined using 5 plants selected from the two central rows of the sampling area. Plots were hand-harvested and cleaned with a mechanical grid with appropriate sieve diameters. A. cruentus was shown to be more suitable to the Central Italy agro-ecological conditions than A. hypochondriacus. The accessions derived from Mexico (PI 477913, PI 576481, PI 643045, PI 643053, and PI 6495079, Guatemala (PI 511719 and Puerto Rico (AMES 5148, had both higher grain yields and a greater stability over the two-year period, with a mean grain production ranging from 2.8 to 3.2 t ha–1. The severe climatic stress in 2012 (high temperatures and aridity, resulted in a 43-60% reduction in seed production compared to that of the previous year. Under these conditions, PI 511719, AMES 26015, AMES 5386, AMES 5148, PI 477913 yielded on average 1.9 t ha–1. Yields of A. hypochondriacus were negligible in both years, probably attributable to greater photoperiod sensitivity, resulting in reduced flowering and delayed maturity. By increasing density up to 60 and 30 plants m–2 for PI 511719 and AMES 5148, respectively, grain production was increased by 55%. As the plant population

  9. High power density carbonate fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Yuh, C.; Johnsen, R.; Doyon, J.; Allen, J. [Energy Research Corp., Danbury, CT (United States)

    1996-12-31

    Carbonate fuel cell is a highly efficient and environmentally clean source of power generation. Many organizations worldwide are actively pursuing the development of the technology. Field demonstration of multi-MW size power plant has been initiated in 1996, a step toward commercialization before the turn of the century, Energy Research Corporation (ERC) is planning to introduce a 2.85MW commercial fuel cell power plant with an efficiency of 58%, which is quite attractive for distributed power generation. However, to further expand competitive edge over alternative systems and to achieve wider market penetration, ERC is exploring advanced carbonate fuel cells having significantly higher power densities. A more compact power plant would also stimulate interest in new markets such as ships and submarines where space limitations exist. The activities focused on reducing cell polarization and internal resistance as well as on advanced thin cell components.

  10. Segmented flow generation by chip reactors for highly parallelized cell cultivation

    Science.gov (United States)

    Grodrian, A.; Metze, J.; Henkel, Thomas; Roth, M.; Kohler, Johann M.

    2002-11-01

    Micro system technology offers convenient tools for the production of handling devices for small liquid volumes which can be used in cell cultivation. Here, a modular system for the rapid generation of cell suspension aliquotes is presented. The system is used to produce and analyze high numbers of strongly separated cultural volumes. Selected clones may be retrieved from the system. Therefore, the principle of segmented flow is applies. Portions of aqueous culture medium containing one cell or very small cell ensembles are separated from each other by a nonmiscible liquid like dodecane or mineral oil. In addition, the oil separates the cultivation droplets from the innerside of the walls of chip channels and capillaries. This way, compatibility problems between cell wall surfaces and the chemical character of technical walls are excluded. The separated cultivation droplets are guided by micro flow transportation in different channel and chamber topologies. The whole system has the character of a serially working cell processing system. The aliquot generation can be speeded up to frequencies of about 30 Hz in each micro channel. That means, that about 105 individual cultural volumes can be produced per hour or about 2 million per day.

  11. Safety of Cultivated Limbal Epithelial Stem Cell Transplantation for Human Corneal Regeneration

    Directory of Open Access Journals (Sweden)

    J. Behaegel

    2017-01-01

    Full Text Available Ex vivo cultivated limbal stem cell transplantation is a promising technique for the treatment of limbal stem cell deficiency. While the results of the clinical trials have been extensively reported since the introduction of the technique in 1997, little has been reported regarding the potential health risks associated with production processes and transplantation techniques. Culture procedures require the use of animal and/or human-derived products, which carry the potential of introducing toxic or infectious agents through contamination with known or unknown additives. Protocols vary widely, and the risks depend on the local institutional methods. Good manufacturing practice and xeno-free culture protocols could reduce potential health risks but are not yet a common practice worldwide. In this review, we focus on the safety of both autologous- and allogeneic-cultivated limbal stem cell transplantation, with respect to culture processes, surgical approaches, and postoperative strategies.

  12. [Cultivation of continuous cell lines on the substrate of carbon nanotubes and the effect of electric stimulation on cell proliferation].

    Science.gov (United States)

    Podcherniaeva, R Ia; Suetina, I A; Mikhaĭlova, G R; Lopatina, O A; Bobrinetskiĭ, I I; Morozov, R A; Seleznev, A S

    2012-01-01

    It was demonstrated that the three studied samples of carbon nanotubes of domestic production fixed on the substrate surface did not have toxic effect and could be used for cell cultivation. A biocompatible conductive coating based on carbon nanotubes and bovine serum albumin was developed. The efficacy of the coating for growing in vitro cell cultures was tested. A device was developed for electric stimulation of the cells. Local electric potential was applied to the cells using nanoscale electrodes. The results of human embryonic fibroblast cultivation in a pulsed electric field on conductive nanocomposite substrates were presented. An 26% increase in the proliferative activity of cells was observed at potentials up to 100 mV.

  13. A SIMPLE WAY OF ACHIEVING A HIGH CELL CONCENTRATION IN RECOMBINANT Escherichia coli CULTIVATION

    Directory of Open Access Journals (Sweden)

    Gombert A.K.

    1997-01-01

    Full Text Available Abstract - A cultivation strategy based on some previous knowledge of the metabolism of Escherichia coli BL21 (DE3 pLysS containing the troponin C gene cloned into plasmid pET was developed and applied through the use of simple fermentation equipment and a feed-forward control strategy in order to achieve a high cell concentration ¾ 92 g l-1 dry cell weight ¾ and a high cell productivity ¾ 3.7 g l-1 h-1.

  14. Nanostructured magnesium increases bone cell density.

    Science.gov (United States)

    Weng, Lucy; Webster, Thomas J

    2012-12-07

    Magnesium has attracted some attention in orthopedics due to its biodegradability and mechanical properties. Since magnesium is an essential natural mineral for bone growth, it can be expected that as a biomaterial, it would support bone formation. However, upon degradation in the body, magnesium releases OH(-) which results in an alkaline pH that can be detrimental to cell density (for example, osteoblasts or bone forming cells). For this reason, modification of magnesium may be necessary to compensate for such detrimental effects to cells. This study created biologically inspired nanoscale surface features on magnesium by soaking magnesium in various concentrations of NaOH (from 1 to 10 N) and for various periods of time (from 10 to 30 min). The results provided the first evidence of increased roughness, surface energy, and consequently greater osteoblast adhesion, after 4 h as well as density up to 7 days on magnesium treated with any concentration of NaOH for any length of time compared to untreated controls. For these reasons, this study suggests that soaking magnesium in NaOH could be an inexpensive, simple and effective manner to promote osteoblast functions for numerous orthopedic applications and, thus, should be further studied.

  15. Comparative study on influence of fetal bovine serum and serum of adult rat on cultivation of newborn rat neural cells

    Directory of Open Access Journals (Sweden)

    Sukach A. N.

    2014-09-01

    Full Text Available Aim. To study the influence of fetal bovine serum and serum of adult rats on behavior of newborn rat isolated neural cells during their cultivation in vitro. Methods. The isolation of neural cells from neonatal rat brain. The determination of the dynamics of cellular monolayer formation. Immunocytochemical staining of cells for β-tubulin III, nestin and vimentin. Results. It has been determined that the addition of serum of adult rats to the cultivation medium creates more favorable conditions for survival, attachment and spread of differentiated, and proliferation of the stem/progenitor neural cells of newborn rats during cultivation in vitro compared with the fetal bovine serum. Conclusions. Using the serum of adult rats is preferable for the cultivation of isolated neural cells of newborn rats compared with the fetal bovine serum.

  16. Plasma modified PLA electrospun membranes for actinorhodin production intensification in Streptomyces coelicolor immobilized-cell cultivations.

    Science.gov (United States)

    Scaffaro, Roberto; Lopresti, Francesco; Sutera, Alberto; Botta, Luigi; Fontana, Rosa Maria; Gallo, Giuseppe

    2017-09-01

    Most of industrially relevant bioproducts are produced by submerged cultivations of actinomycetes. The immobilization of these Gram-positive filamentous bacteria on suitable porous supports may prevent mycelial cell-cell aggregation and pellet formation which usually negatively affect actinomycete submerged cultivations, thus, resulting in an improved biosynthetic capability. In this work, electrospun polylactic acid (PLA) membranes, subjected or not to O 2 -plasma treatment (PLA-plasma), were used as support for immobilized-cell submerged cultivations of Streptomyces coelicolor M145. This strain produces different bioactive compounds, including the blue-pigmented actinorhodin (ACT) and red-pigmented undecylprodigiosin (RED), and constitutes a model for the study of antibiotic-producing actinomycetes. Wet contact angles and X-ray photoelectron spectroscopy analysis confirmed the increased wettability of PLA-plasma due to the formation of polar functional groups such as carboxyl and hydroxyl moieties. Scanning electron microscope observations, carried out at different incubation times, revealed that S. coelicolor immobilized-cells created a dense "biofilm-like" mycelial network on both kinds of PLA membranes. Cultures of S. coelicolor immobilized-cells on PLA or PLA-plasma membranes produced higher biomass (between 1.5 and 2 fold) as well as higher levels of RED and ACT than planktonic cultures. In particular, cultures of immobilized-cells on PLA and PLA-plasma produced comparable levels of RED that were approximatively 4 and 5 fold higher than those produced by planktonic cultures, respectively. In contrast, levels of ACT produced by immobilized-cell cultures on PLA and PLA-plasma were different, being 5 and 10 fold higher than those of planktonic cultures, respectively. Therefore, this is study demonstrated the positive influence of PLA membrane on growth and secondary metabolite production in S. coelicolor and also revealed that O 2 -plasma treated PLA membranes

  17. Effect of monocular deprivation on rabbit neural retinal cell densities

    Directory of Open Access Journals (Sweden)

    Philip Maseghe Mwachaka

    2015-01-01

    Conclusion: In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye.

  18. Effect of explant density and volume of cultivation medium on in-vitro multiplication of blueberry (Vaccinium corymbosum L. varieties "Brigitta" and "Legacy"

    Directory of Open Access Journals (Sweden)

    Mario Rodríguez Beraud

    2015-03-01

    Full Text Available The objective of the investigation was to evaluate the in-vitro multiplication of two varieties of blueberry (Vaccinium corymbosum L., “Brigitta” and “Legacy” in response to five explants densities (5, 10, 15, 20 and 25 and four flask volumes (10, 20, 30 and 40 mL for cultivation. For both varieties the cultivation medium WPM (Woody Plant Medium was used. The experiment was completely randomized with 20 treatments and 12 repetitions per treatment. After 45 days of cultivation we evaluated the height of shoots, number of shoots/explant, number of nodes/shoot and number of shoots/flask. Variety “Brigitta” had highest shoots at higher densities and flask volumes, while variety “Legacy” had the highest average shoot height with intermediate densities and high volumes. Regarding the number of shoots/explant, the volume of the medium had no influence on “Brigitta”, however, higher plant densities affected this parameter. With variety “Legacy” the maximum number of shoots was achieved with lower plant densities and intermediate culture volumes per flask. In relation to the number of nodes per explant "Brigitta had lower numbers as compared to “Legacy”, but with both varieties the number of nodes decresed with smaller volumes of medium in the flasks. For the number of shoots per flask, “Brigitta” responsed best at higher densities exceeding 40 shoots per flask. In contrast, “Legacy” produced maximum results at density of 25 explants in 30 mL of medium. It is concluded that for the optimum multiplication of both varieties the correct selection of both, the planting density and the volume of multiplication medium are important.

  19. Using single cell cultivation system for on-chip monitoring of the interdivision timer in Chlamydomonas reinhardtii cell cycle

    Directory of Open Access Journals (Sweden)

    Soloviev Mikhail

    2010-09-01

    Full Text Available Abstract Regulation of cell cycle progression in changing environments is vital for cell survival and maintenance, and different regulation mechanisms based on cell size and cell cycle time have been proposed. To determine the mechanism of cell cycle regulation in the unicellular green algae Chlamydomonas reinhardtii, we developed an on-chip single-cell cultivation system that allows for the strict control of the extracellular environment. We divided the Chlamydomonas cell cycle into interdivision and division phases on the basis of changes in cell size and found that, regardless of the amount of photosynthetically active radiation (PAR and the extent of illumination, the length of the interdivision phase was inversely proportional to the rate of increase of cell volume. Their product remains constant indicating the existence of an 'interdivision timer'. The length of the division phase, in contrast, remained nearly constant. Cells cultivated under light-dark-light conditions did not divide unless they had grown to twice their initial volume during the first light period. This indicates the existence of a 'commitment sizer'. The ratio of the cell volume at the beginning of the division phase to the initial cell volume determined the number of daughter cells, indicating the existence of a 'mitotic sizer'.

  20. Modeling of hydrodynamics in hollow fiber membrane bioreactor for mammalian cells cultivation

    Directory of Open Access Journals (Sweden)

    N. V. Menshutina

    2016-01-01

    Full Text Available The mathematical modelling in CFD-packages are powerfull instrument for design and calculation of any engineering tasks. CFD-package contains the set of programs that allow to model the different objects behavior based on the mathematical lows. ANSYS Fluent are widely used for modelling of biotechnological and chemical-technological processes. This package is convenient to describe their hydrodynamics. As cell cultivation is one of the actual scientific direction in modern biotechnology ANSYS Fluent was used to create the model of hollow fiber membrane bioreactor. The fibers are hollow cylindrical membrane to be used for cell cultivation. The criterion of process effectiveness for cell growth is full filling of the membrane surface by cells in the bioreactor. While the cell growth the fiber permeability is decreased which effects to feed flow through membrane pores. The specific feature of this process is to ensure such feed flow to deliver the optimal nutrition for the cells on the external membrane surface. The velocity distribution inside the fiber and in all bioreactor as a whole has been calculated based on mass an impulse conservation equations taking into account the mathematical model assumptions. The hydrodynamics analysis in hollow fiber membrane bioreactor is described by the three-dimensional model created in ANSYS Fluent. The specific features of one membrane model are considered and for whole bioreactor too.

  1. Three-dimensional hierarchical cultivation of human skin cells on bio-adaptive hybrid fibers.

    Science.gov (United States)

    Planz, Viktoria; Seif, Salem; Atchison, Jennifer S; Vukosavljevic, Branko; Sparenberg, Lisa; Kroner, Elmar; Windbergs, Maike

    2016-07-11

    The human skin comprises a complex multi-scale layered structure with hierarchical organization of different cells within the extracellular matrix (ECM). This supportive fiber-reinforced structure provides a dynamically changing microenvironment with specific topographical, mechanical and biochemical cell recognition sites to facilitate cell attachment and proliferation. Current advances in developing artificial matrices for cultivation of human cells concentrate on surface functionalizing of biocompatible materials with different biomolecules like growth factors to enhance cell attachment. However, an often neglected aspect for efficient modulation of cell-matrix interactions is posed by the mechanical characteristics of such artificial matrices. To address this issue, we fabricated biocompatible hybrid fibers simulating the complex biomechanical characteristics of native ECM in human skin. Subsequently, we analyzed interactions of such fibers with human skin cells focusing on the identification of key fiber characteristics for optimized cell-matrix interactions. We successfully identified the mediating effect of bio-adaptive elasto-plastic stiffness paired with hydrophilic surface properties as key factors for cell attachment and proliferation, thus elucidating the synergistic role of these parameters to induce cellular responses. Co-cultivation of fibroblasts and keratinocytes on such fiber mats representing the specific cells in dermis and epidermis resulted in a hierarchical organization of dermal and epidermal tissue layers. In addition, terminal differentiation of keratinocytes at the air interface was observed. These findings provide valuable new insights into cell behaviour in three-dimensional structures and cell-material interactions which can be used for rational development of bio-inspired functional materials for advanced biomedical applications.

  2. PCR-activated cell sorting for cultivation-free enrichment and sequencing of rare microbes.

    Directory of Open Access Journals (Sweden)

    Shaun W Lim

    Full Text Available Microbial systems often exhibit staggering diversity, making the study of rare, interesting species challenging. For example, metagenomic analyses of mixed-cell populations are often dominated by the sequences of the most abundant organisms, while those of rare microbes are detected only at low levels, if at all. To overcome this, selective cultivation or fluorescence-activated cell sorting (FACS can be used to enrich for the target species prior to sequence analysis; however, since most microbes cannot be grown in the lab, cultivation strategies often fail, while cell sorting requires techniques to uniquely label the cell type of interest, which is often not possible with uncultivable microbes. Here, we introduce a culture-independent strategy for sorting microbial cells based on genomic content, which we term PCR-activated cell sorting (PACS. This technology, which utilizes the power of droplet-based microfluidics, is similar to FACS in that it uses a fluorescent signal to uniquely identify and sort target species. However, PACS differs importantly from FACS in that the signal is generated by performing PCR assays on the cells in microfluidic droplets, allowing target cells to be identified with high specificity with suitable design of PCR primers and TaqMan probes. The PACS assay is general, requires minimal optimization and, unlike antibody methods, can be developed without access to microbial antigens. Compared to non-specific methods in which cells are sorted based on size, granularity, or the ability to take up dye, PACS enables genetic sequence-specific sorting and recovery of the cell genomes. In addition to sorting microbes, PACS can be applied to eukaryotic cells, viruses, and naked nucleic acids.

  3. Scalable cultivation of human pluripotent stem cells on chemically-defined surfaces

    Science.gov (United States)

    Hsiung, Michael Chi-Wei

    Human stem cells (SCs) are classified as self-renewing cells possessing great ability in therapeutic applications due of their ability to differentiate along any major cell lineage in the human body. Despite their restorative potential, widespread use of SCs is hampered by strenuous control issues. Along with the need for strict xeno-free environments to sustain growth in culture, current methods for growing human pluripotent stem cells (hPSCs) rely on platforms which impede large-scale cultivation and therapeutic delivery. Hence, any progress towards development of large-scale culture systems is severely hindered. In a concentrated effort to develop a scheme that can serve as a model precursor for large scale SC propagation in clinical use, we have explored methods for cultivating hPSCs on completely defined surfaces. We discuss novel approaches with the potential to go beyond the limitations presented by current methods. In particular, we studied the cultivation of human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) on surface which underwent synthetic or chemical modification. Current methods for hPSCs rely on animal-based extracellular matrices (ECMs) such as mouse embryonic fibroblasts (MEFs) or feeders and murine sacoma cell-derived substrates to facilitate their growth. While these layers or coatings can be used to maximize the output of hPSC production, they cannot be considered for clinical use because they risk introducing foreign pathogens into culture. We have identified and developed conditions for a completely defined xeno-free substrate used for culturing hPSCs. By utilizing coupling chemistry, we can functionalize ester groups on a given surface and conjugate synthetic peptides containing the arginine-glycine-aspartic acid (RGD) motif, known for their role in cell adhesion. This method offers advantages over traditional hPSC culture by keeping the modified substrata free of xenogenic response and can be scaled up in

  4. Recent Advances in Outdoor High-Density Cultivation of Novelty Micro-Algae Strain with High Content of Lipids

    OpenAIRE

    Kaštánek, Petr

    2012-01-01

    The objective of the study was the pilot plant examination of a newly developed integrated process for autotrophic cultivation of useful micro-algae. The process utilizes waste carbon dioxide as a source of carbon and yields simultaneously products that can be utilized in food and cosmetic industries, turned into biodiesel and/or used as a supplement in animal feed. At present, the cultivation of micro-algae merely for the production of biofuels is not economically viable. In the proposed pr...

  5. Cell growth stimulating effect of Ganoderma lucidum spores and their potential application for Chinese hamster ovary K1 cell cultivation.

    Science.gov (United States)

    Li, Ding; Zhong, Qi; Liu, Tingting; Wang, Jufang

    2016-06-01

    In this work, water-soluble extracts of Ganoderma lucidum spores (Gls), a Chinese medicinal herb that possesses cell growth stimulating function, were found to be an effective growth factor for Chinese hamster ovary (CHO) cell cultivation. The Gls extract was prepared and supplemented to CHO K1 cell culture media with various serum levels. Our results obtained from both the static culture and the spinner-flask suspension culture showed that use of small-amount Gls extract effectively promoted cell growth and suppressed cell apoptosis induced by serum deprivation with normal cell cycle maintained in a low-serum medium. The low-serum medium containing 1 % (v/v) fetal bovine serum (FBS) and 0.01 % (w/v) Gls extract showed a comparable performance on both cell growth and fusion protein productivity with the conventional CHO culture medium containing 10 % (v/v) FBS and a commercial serum-free medium. This is the first study of the potential of Gls extracts for use as an alternative cell growth factor and nutrient for CHO cells. The findings have presented a new approach to economic cultivation of CHO cells for therapeutic protein production.

  6. Microfluidic devices for stem-cell cultivation, differentiation and toxicity testing

    Science.gov (United States)

    Becker, Holger; Hansen-Hagge, Thomas; Kurtz, Andreas; Mrowka, Ralf; Wölfl, Stefan; Gärtner, Claudia

    2017-02-01

    The development of new drugs is time-consuming, extremely expensive and often promising drug candidates fail in late stages of the development process due to the lack of suitable tools to either predict toxicological effects or to test drug candidates in physiologically relevant environments prior to clinical tests. We therefore try to develop diagnostic multiorgan microfluidic chips based on patient specific induced pluripotent stem cell (iPS) technology to explore liver dependent toxic effects of drugs on individual human tissues such as liver or kidney cells. Based initially on standardized microfluidic modules for cell culture, we have developed integrated microfluidic devices which contain different chambers for cell/tissue cultivation. The devices are manufactured using injection molding of thermoplastic polymers such as polystyrene or cyclo-olefin polymer. In the project, suitable surface modification methods of the used materials had to be explored. We have been able to successfully demonstrate the seeding, cultivation and further differentiation of modified iPS, as shown by the use of differentiation markers, thus providing a suitable platform for toxicity testing and potential tissue-tissue interactions.

  7. Cheese whey-induced high-cell-density production of recombinant proteins in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Neubauer Peter

    2003-04-01

    Full Text Available Abstract Background Use of lactose-rich concentrates from dairy processes for the induction of recombinant gene's expression has not received much attention although they are interesting low cost substrates for production of recombinant enzymes. Applicability of dairy waste for induction of recombinant genes in Escherichia coli was studied. Clones expressing Lactobacillus phage muramidase and Lactobacillus alcohol dehydrogenase were used for the experiments. Results Shake flask cultivations in mineral salt medium showed that cheese whey or deproteinised whey induced gene expression as efficiently as IPTG (isopropyl-β-D-thiogalactopyranoside or pure lactose. Addition of yeast extract or proteolytically degraded whey proteins did not improve the recombinant protein yield. In contrast, addition of yeast extract to the well-balanced mineral salt medium decreased the product yield. Feeding with glycerol provided sufficient amount of easily assimilable carbon source during the induction period without preventing lactose intake and induction by lactose. High-cell-density fed-batch cultivations showed that product yields comparable to IPTG-induction can be achieved by feeding bacteria with a mixture of glycerol and concentrated whey permeate during the induction. Conclusion Whey and concentrated whey permeate can be applied as an alternative inducer in recombinant high-cell-density fed-batch fermentations. The yield of the recombinant product was comparable to fermentations induced by IPTG. In low-cell-density shake flask experiments the yield was higher with whey or whey permeate than with IPTG.

  8. Thermoforming techniques for manufacturing porous scaffolds for application in 3D cell cultivation.

    Science.gov (United States)

    Borowiec, Justyna; Hampl, Jörg; Gebinoga, Michael; Elsarnagawy, Tarek; Elnakady, Yasser A; Fouad, Hassan; Almajhadi, Fahd; Fernekorn, Uta; Weise, Frank; Singh, Sukhdeep; Elsarnagawy, Dief; Schober, Andreas

    2015-04-01

    Within the scientific community, there is an increasing demand to apply advanced cell cultivation substrates with increased physiological functionalities for studying spatially defined cellular interactions. Porous polymeric scaffolds are utilized for mimicking an organ-like structure or engineering complex tissues and have become a key element for three-dimensional (3D) cell cultivation in the meantime. As a consequence, efficient 3D scaffold fabrication methods play an important role in modern biotechnology. Here, we present a novel thermoforming procedure for manufacturing porous 3D scaffolds from permeable materials. We address the issue of precise thermoforming of porous polymer foils by using multilayer polymer thermoforming technology. This technology offers a new method for structuring porous polymer foils that are otherwise available for non-porous polymers only. We successfully manufactured 3D scaffolds from solvent casted and phase separated polylactic acid (PLA) foils and investigated their biocompatibility and basic cellular performance. The HepG2 cell culture in PLA scaffold has shown enhanced albumin secretion rate in comparison to a previously reported polycarbonate based scaffold with similar geometry. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Microalgae cultivation in a tubular bioreactor and utilization of their cells

    Science.gov (United States)

    Koyu, Hon-Nami; Shunji, Kunito

    1998-03-01

    In this study on the possiblities of microalgae technology as an option for CO2 mitigation, many microalgae were isolated from seawater. Some species of the isolates, Chlamydomonas sp. strain YA-SH-1, which accumulates starch in cells under light and ferment ethanol in dark and anaerobic condition, was grown outdoors by using 50-L tubular bioreactors in batch cultivation and harvested. Using these cells, the performance of ethanol production was examined quantitatively in a 0.5-L scale fermentor. Another species, Tetraselmis sp. strain Tt-1, was cultivated in a semi-batch manner by a similar type of tubular bioreactor indoors and examined for its utilization. Tests showed these cells could be used as partial substitute for wood and kenaf pulp for processing into paper. With the idea of making microalgae produce cellulose by genetic engineering in their minds, the authors studied the structure of bacterial cellulose synthase genes and the low temperature-induced, reversible flocculation in a thermophilic blue green alga (Cyanobacterium), Synechocystis vulcanus in order to examine the feasibility of using these genes as gene source and the cynanobacterium as host.

  10. Cytotoxic activity of ethanolic extracts of Eleutherococcus species cultivated in Poland on HL60 leukemia cell line

    OpenAIRE

    Zaluski Daniel; Smolarz Helena Danuta; Bogucka-Kocka Anna

    2014-01-01

    The Eleutherococcus species including 40 species native to Asia are medicinal plants widely used in traditional medicine. Some of these species are cultivated at the botanical gardens in Europe. On the basis on our earlier studies it was concluded that the extracts of analyzed species act as antioxidants, inhibitors of MMPs, and cytotoxic against Jurkat 45 leukemia cell line. In this study, the anti-leukemic potential of roots and leaves from six Eleutherococcus species cultivated in Poland w...

  11. Impact of cultivation conditions on N-glycosylation of influenza virus a hemagglutinin produced in MDCK cell culture.

    Science.gov (United States)

    Rödig, Jana Verena; Rapp, Erdmann; Bohne, Jana; Kampe, Michael; Kaffka, Helene; Bock, Andreas; Genzel, Yvonne; Reichl, Udo

    2013-06-01

    Manufacturers worldwide produce influenza vaccines in different host systems. So far, either fertilized chicken eggs or mammalian cell lines are used. In all these vaccines, hemagglutinin (HA) and neuraminidase are the major components. Both are highly abundant glycoproteins in the viral envelope, and particularly HA is able to induce a strong and protective immune response. The quality characteristics of glycoproteins, such as specific activity, antigenicity, immunogenicity, binding avidity, and receptor-binding specificity can strongly depend on changes or differences in their glycosylation pattern (potential N-glycosylation occupancy as well as glycan composition). In this study, capillary gel electrophoresis with laser-induced fluorescence detection (CGE-LIF) based glycoanalysis (N-glycan fingerprinting) was used to determine the impact of cultivation conditions on the HA N-glycosylation pattern of Madin-Darby canine kidney (MDCK) cell-derived influenza virus A PR/8/34 (H1N1). We found that adaptation of adherent cells to serum-free growth has only a minor impact on the HA N-glycosylation pattern. Only relative abundances of N-glycan structures are affected. In contrast, host cell adaptation to serum-free suspension growth resulted in significant changes in the HA N-glycosylation pattern regarding the presence of specific N-glycans as well as their abundance. Further controls such as different suppliers for influenza virus A PR/8/34 (H1N1) seed strains, different cultivation scales and vessels in standard or high cell density mode, different virus production media varying in either composition or trypsin activity, different temperatures during virus replication and finally, the impact of β-propiolactone inactivation resulted-at best-only in minor changes in the relative N-glycan structure abundances of the HA N-glycosylation pattern. Surprisingly, these results demonstrate a rather stable HA N-glycosylation pattern despite various (significant) changes in

  12. A Strategy for Cultivation of Retargeted Oncolytic Herpes Simplex Viruses in Non-cancer Cells.

    Science.gov (United States)

    Leoni, Valerio; Gatta, Valentina; Casiraghi, Costanza; Nicosia, Alfredo; Petrovic, Biljana; Campadelli-Fiume, Gabriella

    2017-05-15

    The oncolytic herpes simplex virus (HSV) that has been approved for clinical practice and those HSVs in clinical trials are attenuated viruses, often with the neurovirulence gene γ 1 34.5 and additional genes deleted. One strategy to engineer nonattenuated oncolytic HSVs consists of retargeting the viral tropism to a cancer-specific receptor of choice, exemplified by HER2 (human epidermal growth factor receptor 2), which is present in breast, ovary, and other cancers, and in detargeting from the natural receptors. Because the HER2-retargeted HSVs strictly depend on this receptor for infection, the viruses employed in preclinical studies were cultivated in HER2-positive cancer cells. The production of clinical-grade viruses destined for humans should avoid the use of cancer cells. Here, we engineered the R-213 recombinant, by insertion of a 20-amino-acid (aa) short peptide (named GCN4) in the gH of R-LM113; this recombinant was retargeted to HER2 through insertion in gD of a single-chain antibody (scFv) to HER2. Next, we generated a Vero cell line expressing an artificial receptor (GCN4R) whose N terminus consists of an scFv to GCN4 and therefore is capable of interacting with GCN4 present in gH of R-213. R-213 replicated as well as R-LM113 in SK-OV-3 cells, implying that addition of the GCN4 peptide was not detrimental to gH. R-213 grew to relatively high titers in Vero-GCN4R cells, efficiently spread from cell to cell, and killed both Vero-GCN4R and SK-OV-3 cells, as expected for an oncolytic virus. Altogether, Vero-GCN4R cells represent an efficient system for cultivation of retargeted oncolytic HSVs in non-cancer cells. IMPORTANCE There is growing interest in viruses as oncolytic agents, which can be administered in combination with immunotherapeutic compounds, including immune checkpoint inhibitors. The oncolytic HSV approved for clinical practice and those in clinical trials are attenuated viruses. An alternative to attenuation is a cancer specificity

  13. Efficacy of cultivated corneal epithelial stem cells for ocular surface reconstruction

    Directory of Open Access Journals (Sweden)

    Prabhasawat P

    2012-09-01

    Full Text Available Pinnita Prabhasawat,1 Pattama Ekpo,2 Mongkol Uiprasertkul,3 Suksri Chotikavanich,1 Nattaporn Tesavibul11Department of Ophthalmology, 2Department of Immunology, 3Department of Pathology, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, ThailandPurpose: To investigate the clinical outcomes of cultivated corneal limbal epithelial transplantation (CLET using human amniotic membrane for corneal limbal stem-cell deficiency.Methods: Prospective, noncomparative case series. Eighteen patients (19 eyes with severe ocular surface diseases were chosen to undergo CLET using human amniotic membrane. Twelve eyes received auto-CLET, and seven eyes received allo-CLET. Clinical outcomes of corneal surface epithelialization, conjunctivalization, inflammation, visual acuity, graft status, and complications were observed.Results: Corneal epithelium cultivated on amniotic membrane (two to four layers was positive for molecular markers p63, ABCG2, CK3, and CK12. The mean patient age was 44.7 ± 15.2 years. A successful clinical outcome, defined as corneal epithelialization without central conjunctivalization or severe inflammation, was obtained in 14 (73.7% of 19 eyes (mean follow-up 26.1 ± 13.5 months; range 6–47. A histopathologic success, defined as absence of goblet cells at the central cornea, was achieved in 12 (63.2% eyes. Clinical failures occurred in five (26.3% of 19 eyes, and histopathologic failures occurred in seven (36.8% of 19 eyes. Survival analysis at 1 year showed that the clinical success rate was 77.9% and the pathological success rate was 72.3%. Fourteen of 19 (73.7% eyes had visual acuity improvements after CLET. Six cases underwent penetrating keratoplasty; five of these grafts remained clear after 20.4 ± 6.9 months (range, 12–31 of follow-up. Complications included infectious keratitis (three cases and recurrent symblepharon (one case. All complicated cases had lid abnormalities. Factors affecting the final clinical

  14. Absence of micronucleus formation in CHO-K1 cells cultivated in platelet lysate enriched medium.

    Science.gov (United States)

    Bernardi, Martina; Adami, Valentina; Albiero, Elena; Madeo, Domenico; Rodeghiero, Francesco; Astori, Giuseppe

    2014-03-01

    Human platelet lysate (PL) represents an effective substitute of fetal bovine serum (FBS) for mesenchymal stromal cell (MSC) cultivation. Compared to FBS, PL favors MSC proliferation significantly shortening the population doubling time and avoiding the risks related to the use of animal derivatives. Growth factors contained in the platelets are released upon platelet disruption following freezing/thawing cycles or as we have recently described by using ultrasound. We have investigated whether the increased cell proliferation achieved by using PL could induce mitotic stress and whether the potential formation of free radicals during PL production by ultrasound could cause chromosomal instability in mammalian cells. We have applied an image analysis assisted high content screening (HCS) in vitro micronucleus assay in the Chinese Hamster Ovarian K1 (CHO-K1) rodent mammalian cell line. PL was produced by sonication; for the micronucleus assay, CHO-K1 cells were exposed to increasing concentrations of PL. Cytokinesis was blocked by cytochalasin B, nuclei were stained with bisbenzimide and images were acquired and analyzed automatically using an HCS system, both with a 20× and a 10× objective. Our results suggest that growth stimulus induced by the use of PL did not significantly increase micronucleus formation in CHO-K1 cells compared to negative control. Micronucleus testing in conjunction with HCS could represent a valid tool to evaluate the safety of ancillary materials used in the production of cell-based medicinal products. Copyright © 2013 Elsevier GmbH. All rights reserved.

  15. Cultivation of Human Microvascular Endothelial Cells on Topographical Substrates to Mimic the Human Corneal Endothelium

    Directory of Open Access Journals (Sweden)

    Jie Shi Chua

    2013-03-01

    Full Text Available Human corneal endothelial cells have a limited ability to replicate in vivo and in vitro. Allograft transplantation becomes necessary when an accident or trauma results in excessive cell loss. The reconstruction of the cornea endothelium using autologous cell sources is a promising alternative option for therapeutic or in vitro drug testing applications. The native corneal endothelium rests on the Descemet’s membrane, which has nanotopographies of fibers and pores. The use of synthetic topographies mimics the native environment, and it is hypothesized that this can direct the behavior and growth of human microvascular endothelial cells (HMVECs to resemble the corneal endothelium. In this study, HMVECs are cultivated on substrates with micron and nano-scaled pillar and well topographies. Closely packed HMVEC monolayers with polygonal cells and well-developed tight junctions were formed on the topographical substrates. Sodium/potassium (Na+/K+ adenine triphosphatase (ATPase expression was enhanced on the microwells substrate, which also promotes microvilli formation, while more hexagonal-like cells are found on the micropillars samples. The data obtained suggests that the use of optimized surface patterning, in particular, the microtopographies, can induce HMVECs to adopt a more corneal endothelium-like morphology with similar barrier and pump functions. The mechanism involved in cell contact guidance by the specific topographical features will be of interest for future studies.

  16. A Protocol for Isolation and Enriched Monolayer Cultivation of Neural Precursor Cells from Mouse Dentate Gyrus

    OpenAIRE

    Babu, Harish; Claasen, Jan-Hendrik; Kannan, Suresh; Rünker, Annette E.; Palmer, Theo; Kempermann, Gerd

    2011-01-01

    In vitro assays are valuable tools to study the characteristics of adult neural precursor cells under controlled conditions with a defined set of parameters. We here present a detailed protocol based on our previous original publication (Babu et al., 2007) to isolate neural precursor cells from the hippocampus of adult mice and maintain and propagate them as adherent monolayer cultures. The strategy is based on the use of Percoll density gradient centrifugation to enrich precursor cells from ...

  17. Comparison of culture media for ex vivo cultivation of limbal epithelial progenitor cells.

    Science.gov (United States)

    Loureiro, Renata Ruoco; Cristovam, Priscila Cardoso; Martins, Caio Marques; Covre, Joyce Luciana; Sobrinho, Juliana Aparecida; Ricardo, José Reinaldo da Silva; Hazarbassanov, Rossen Myhailov; Höfling-Lima, Ana Luisa; Belfort, Rubens; Nishi, Mauro; Gomes, José Álvaro Pereira

    2013-01-01

    To compare the effectiveness of three culture media for growth, proliferation, differentiation, and viability of ex vivo cultured limbal epithelial progenitor cells. Limbal epithelial progenitor cell cultures were established from ten human corneal rims and grew on plastic wells in three culture media: supplemental hormonal epithelial medium (SHEM), keratinocyte serum-free medium (KSFM), and Epilife. The performance of culturing limbal epithelial progenitor cells in each medium was evaluated according to the following parameters: growth area of epithelial migration; immunocytochemistry for adenosine 5'-triphosphate-binding cassette member 2 (ABCG2), p63, Ki67, cytokeratin 3 (CK3), and vimentin (VMT) and real-time reverse transcription polymerase chain reaction (RT-PCR) for CK3, ABCG2, and p63, and cell viability using Hoechst staining. Limbal epithelial progenitor cells cultivated in SHEM showed a tendency to faster migration, compared to KSFM and Epilife. Immunocytochemical analysis showed that proliferated cells in the SHEM had lower expression for markers related to progenitor epithelial cells (ABCG2) and putative progenitor cells (p63), and a higher percentage of positive cells for differentiated epithelium (CK3) when compared to KSFM and Epilife. In PCR analysis, ABCG2 expression was statistically higher for Epilife compared to SHEM. Expression of p63 was statistically higher for Epilife compared to SHEM and KSFM. However, CK3 expression was statistically lower for KSFM compared to SHEM. Based on our findings, we concluded that cells cultured in KSFM and Epilife media presented a higher percentage of limbal epithelial progenitor cells, compared to SHEM.

  18. Relationships between blood cell counts and the density of malaria ...

    African Journals Online (AJOL)

    The findings depicted a negative correlation between parasite load and haemoglobin concentration [Hb], mean cell volume (MCV), and mean cell haemoglobin (MCH); a positive correlation of parasite density with white blood cell counts (WBC), red blood cell counts (RBC), and the differential white blood cell counts ...

  19. Biodegradation and Osteosarcoma Cell Cultivation on Poly(aspartic acid) Based Hydrogels.

    Science.gov (United States)

    Juriga, Dávid; Nagy, Krisztina; Jedlovszky-Hajdú, Angéla; Perczel-Kovách, Katalin; Chen, Yong Mei; Varga, Gábor; Zrínyi, Miklós

    2016-09-14

    Development of novel biodegradable and biocompatible scaffold materials with optimal characteristics is important for both preclinical and clinical applications. The aim of the present study was to analyze the biodegradability of poly(aspartic acid)-based hydrogels, and to test their usability as scaffolds for MG-63 osteoblast-like cells. Poly(aspartic acid) was fabricated from poly(succinimide) and hydrogels were prepared using natural amines as cross-linkers (diaminobutane and cystamine). Disulfide bridges were cleaved to thiol groups and the polymer backbone was further modified with RGD sequence. Biodegradability of the hydrogels was evaluated by experiments on the base of enzymes and cell culture medium. Poly(aspartic acid) hydrogels possessing only disulfide bridges as cross-links proved to be degradable by collagenase I. The MG-63 cells showed healthy, fibroblast-like morphology on the double cross-linked and RGD modified hydrogels. Thiolated poly(aspartic acid) based hydrogels provide ideal conditions for adhesion, survival, proliferation, and migration of osteoblast-like cells. The highest viability was found on the thiolated PASP gels while the RGD motif had influence on compacted cluster formation of the cells. These biodegradable and biocompatible poly(aspartic acid)-based hydrogels are promising scaffolds for cell cultivation.

  20. Effect of different sowing densities in mixed cultivation of blue lupin (Lupinus angustifolius) with spring crops on yield and quality

    OpenAIRE

    Böhm, Herwart; Bramm, Andreas; Aulrich, Karen; Rühl, Gerhard

    2008-01-01

    The aim of this investigation was to optimise the yield percentage of blue lupins in mixed cropping systems. Field experiments were conducted at two locations in Northern Germany (Institute of Organic Farming near Hamburg and Institute of Plant and Soil Science at Braunschweig). Two types of blue lupins, the determinate and branched type were cultivated together with spring barley, spring wheat or fodder peas in different seeding ratios (SR): 50% : 50%, 62,5% : 37,5 % and 75% : 25% of the res...

  1. Production of nattokinase by high cell density fed-batch culture of Bacillus subtilis.

    Science.gov (United States)

    Kwon, Eun-Yeong; Kim, Kyung Mi; Kim, Mi Kyoung; Lee, In Young; Kim, Beom Soo

    2011-09-01

    Bacillus subtilis was cultivated to high cell density for nattokinase production by pH-stat fed-batch culture. A concentrated mixture solution of glucose and peptone was automatically added by acid-supplying pump when culture pH rose above high limit. Effect of the ratio of glucose to peptone in feeding solution was investigated on cell growth and nattokinase production by changing the ratio from 0.2 to 5 g glucose/g peptone. The highest cell concentration was 77 g/L when the ratio was 0.2 g glucose/g peptone. Cell concentration decreased with increasing the ratio of glucose to peptone in feeding solution, while the optimum condition existed for nattokinase production. The highest nattokinase activity was 14,500 unit/mL at a ratio of 0.33 g glucose/g peptone, which was 4.3 times higher than that in batch culture.

  2. Cultivation of mammalian cells using a single-use pneumatic bioreactor system.

    Science.gov (United States)

    Obom, Kristina M; Cummings, Patrick J; Ciafardoni, Janelle A; Hashimura, Yasunori; Giroux, Daniel

    2014-10-10

    Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized medicine innovations. However, translating these advances into therapeutic applications will require in vitro systems that allow for robust, flexible, and cost effective bioreactor systems. There are several bioreactor systems currently utilized in research and commercial settings; however, many of these systems are not optimal for establishing, expanding, and monitoring the growth of different cell types. The culture parameters most challenging to control in these systems include, minimizing hydrodynamic shear, preventing nutrient gradient formation, establishing uniform culture medium aeration, preventing microbial contamination, and monitoring and adjusting culture conditions in real-time. Using a pneumatic single-use bioreactor system, we demonstrate the assembly and operation of this novel bioreactor for mammalian cells grown on micro-carriers. This bioreactor system eliminates many of the challenges associated with currently available systems by minimizing hydrodynamic shear and nutrient gradient formation, and allowing for uniform culture medium aeration. Moreover, the bioreactor's software allows for remote real-time monitoring and adjusting of the bioreactor run parameters. This bioreactor system also has tremendous potential for scale-up of adherent and suspension mammalian cells for production of a variety therapeutic proteins, monoclonal antibodies, stem cells, biosimilars, and vaccines.

  3. Cultivation of Rickettsia amblyommii in tick cells, prevalence in Florida lone star ticks (Amblyomma americanum).

    Science.gov (United States)

    Sayler, Katherine A; Wamsley, Heather L; Pate, Melanie; Barbet, Anthony F; Alleman, A Rick

    2014-06-13

    Rickettsia amblyommii is a bacterium in the spotted fever group of organisms associated with the lone star tick (LST), Amblyomma americanum. The LST is the most commonly reported tick to parasitize humans in the southeastern US. Within this geographic region, there have been suspected cases of Rocky Mountain spotted fever (RMSF) where the causative agent, R. rickettsii, was not identified in the local tick population. In these areas, patients with clinical signs of RMSF had low or no detectable antibodies to R. rickettsii, resulting in an inability to confirm a diagnosis. R. amblyommii was cultivated from host-seeking LSTs trapped in Central Florida and propagated in ISE6 (Ixodes scapularis) and AAE2 (A. americanum) cells. Quantitative PCR targeting the 17-kD gene of Rickettsia spp. identified the genus of the organism in culture. Variable regions of groEL, gtlA and rompA genes were amplified and sequenced to confirm the species. The prevalence of R. amblyommii in LSTs within the geographic region was determined by qPCR followed by conventional PCR and direct sequencing. Analyses of amplified sequences from the cultured organism were 100% homologous to R. amblyommii. The overall prevalence of Rickettsia spp. in the local population of LSTs was 57.1% and rompA sequence analysis identified only R. amblyommii in LSTs. A Florida strain of R. amblyommii was successfully cultivated in two tick cell lines. Further evaluation of the new strain and comparisons to the other geographic strains is needed. The prevalence of this SFG organism in the tick population warrants further investigation into the organism's ability to cause clinical disease in mammalian species.

  4. Two Cases of Non-Small Cell Lung Cancer Treated with Intravenous Cultivated Wild Ginseng Pharmacopuncture

    Directory of Open Access Journals (Sweden)

    Sun-Hwi Bang

    2008-06-01

    Full Text Available Objectives : To investigate the therapeutic effects of intravenous cultivated wild ginseng(Panax ginseng C.A. Meyer pharmacopuncture(CWGP in treating patients with non-small cell lung cancer(NSCLC. Design : Prospective case series. Setting : This study was conducted at the East-West Cancer Center of Dunsan Oriental Hospital, Daejeon University. Patients : Two non-small cell lung cancer patients. Intervention : Two non-small cell lung cancer patients were injected CWGP(20mL/day mixed with 0.9% normal saline(100mL intravenously. Each patient received a total of 16 and 9 cycles, respectively. One cycle is composed of 14 days. Outcome Measures : The effect of intravenous CWGP was measured by scanning with computed tomography(CT after every 2 cycle and Positron emission tomography- computed tomography(PET/CT after every 6 cycles. Response and progression was evaluated using the Response Evaluation Criteria in Solid Tumors(RECIST Committee classification of complete response(CR, partial response(PR, progressive disease(PD and stable disease(SD. Results : They were treated with intravenous CWGP for 8 and 5 months respectively. time later, each tumor remains stable disease(SD Conclusion : These cases may give us a possibility that intravenous CWGP offers potential benefits for non-small cell lung cancer patients.

  5. Co-cultivation of murine BMDCs with 67NR mouse mammary carcinoma cells give rise to highly drug resistant cells

    Directory of Open Access Journals (Sweden)

    Zänker Kurt S

    2011-06-01

    Full Text Available Abstract Background Tumor tissue resembles chronically inflamed tissue. Since chronic inflammatory conditions are a strong stimulus for bone marrow-derived cells (BMDCs it can be assumed that recruitment of BMDCs into cancer tissue should be a common phenomenon. Several data have outlined that BMDC can influence tumor growth and metastasis, e.g., by inducing a paracrine acting feedback loop in tumor cells. Likewise, cell fusion and horizontal gene transfer are further mechanisms how BMDCs can trigger tumor progression. Results Hygromycin resistant murine 67NR-Hyg mammary carcinoma cells were co-cultivated with puromycin resistant murine BMDCs from Tg(GFPU5Nagy/J mice. Isolation of hygromycin/puromycin resistant mBMDC/67NR-Hyg cell clones was performed by a dual drug selection procedure. PCR analysis revealed an overlap of parental markers in mBMDC/67NR-Hyg cell clones, suggesting that dual resistant cells originated by cell fusion. By contrast, both STR and SNP data analysis indicated that only parental 67NR-Hyg alleles were found in mBMDC/67NR-Hyg cell clones favoring horizontal gene transfer as the mode of origin. RealTime-PCR-array analysis showed a marked up-regulation of Abcb1a and Abcb1b ABC multidrug transporters in mBMDC/67NR-Hyg clones, which was verified by Western Blot analysis. Moreover, the markedly increased Abcb1a/Abcb1b expression was correlated to an efficient Rhodamine 123 efflux, which was completely inhibited by verapamil, a well-known Abcb1a/Abcb1b inhibitor. Likewise, mBMDCs/67NR-Hyg clones revealed a marked resistance towards chemotherapeutic drugs including 17-DMAG, doxorubicin, etoposide and paclitaxel. In accordance to Rhodamine 123 efflux data, chemotherapeutic drug resistance of mBMDC/67NR-Hyg cells was impaired by verapamil mediated blockage of Abc1a/Abcb1b multidrug transporter function. Conclusion Co-cultivation of mBMDCs and mouse 67NR-Hyg mammary carcinoma cells gave rise to highly drug resistant cells. Even

  6. Application of nano TiO2 modified hollow fiber membranes in algal membrane bioreactors for high-density algae cultivation and wastewater polishing.

    Science.gov (United States)

    Hu, Weiming; Yin, Jun; Deng, Baolin; Hu, Zhiqiang

    2015-10-01

    Polyvinylidene fluoride (PVDF) hollow fiber membranes with nano-TiO2 (5% of PVDF by mass, average size = 25 nm) additives were fabricated and applied for high-density algae (Chlorella vulgaris) cultivation. At the average light intensity of 121 μmol/m(2)/s, the algal membrane bioreactors (A-MBR) operated at a hydraulic retention time of 0.5d and an average solids retention time of 25d had an average algae biomass concentration of 2350 ± 74 mg/L (in COD units) and algal biomass production rate of 6.5 ± 0.1g/m(2)/d. The A-MBRs removed an average of 78% of phosphorus from the wastewater at the initial total phosphorus concentrations ranging from 3.5 to 8.6 mg/L. The nano TiO2-embedded membranes had improved surface hydrophilicity with its total resistance about 50% lower than that of the control. This study demonstrated that PVDF/TiO2 nanocomposite membranes had a better antifouling property for high-density algae cultivation and wastewater polishing. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Dynamic modelling of high biomass density cultivation and biohydrogen production in different scales of flat plate photobioreactors.

    Science.gov (United States)

    Zhang, Dongda; Dechatiwongse, Pongsathorn; Del Rio-Chanona, Ehecatl Antonio; Maitland, Geoffrey C; Hellgardt, Klaus; Vassiliadis, Vassilios S

    2015-12-01

    This paper investigates the scaling-up of cyanobacterial biomass cultivation and biohydrogen production from laboratory to industrial scale. Two main aspects are investigated and presented, which to the best of our knowledge have never been addressed, namely the construction of an accurate dynamic model to simulate cyanobacterial photo-heterotrophic growth and biohydrogen production and the prediction of the maximum biomass and hydrogen production in different scales of photobioreactors. To achieve the current goals, experimental data obtained from a laboratory experimental setup are fitted by a dynamic model. Based on the current model, two key original findings are made in this work. First, it is found that selecting low-chlorophyll mutants is an efficient way to increase both biomass concentration and hydrogen production particularly in a large scale photobioreactor. Second, the current work proposes that the width of industrial scale photobioreactors should not exceed 0.20 m for biomass cultivation and 0.05 m for biohydrogen production, as severe light attenuation can be induced in the reactor beyond this threshold. © 2015 Wiley Periodicals, Inc.

  8. In vitro cultivation of malignant lymphoblasts of transplantable Mouse Lymphosarcoma MB (T 86157) without typical mesenchyme cells

    NARCIS (Netherlands)

    Bruyn, de Willemina M.

    1949-01-01

    Previous investigations (see Literature) by means of tissue culture methods have shown that the malignant lymphoblasts of mouse lymphosarcoma MB (T 86157) can be cultivated indefinitely when in the presence of actively growing mesenchyme cells. Under the cultural conditions provided, which included

  9. Producing biodiesel from cotton seed oil using Rhizopus oryzae ATTC #34612 whole cell biocatalysts: Culture media and cultivation period optimization

    Science.gov (United States)

    The effect of culture medium composition and cultivation time on biodiesel production by Rhizopus oryzae ATCC #34612 whole cell catalysts, immobilized on novel rigid polyethylene biomass supports, was investigated. Supplementation of the medium with carbon sources led to higher lipase activity and i...

  10. Very high cell density perfusion of CHO cells anchored in a non-woven matrix-based bioreactor.

    Science.gov (United States)

    Zhang, Ye; Stobbe, Per; Silvander, Christian Orrego; Chotteau, Véronique

    2015-11-10

    Recombinant Chinese Hamster Ovary (CHO) cells producing IgG monoclonal antibody were cultivated in a novel perfusion culture system CellTank, integrating the bioreactor and the cell retention function. In this system, the cells were harbored in a non-woven polyester matrix perfused by the culture medium and immersed in a reservoir. Although adapted to suspension, the CHO cells stayed entrapped in the matrix. The cell-free medium was efficiently circulated from the reservoir into- and through the matrix by a centrifugal pump placed at the bottom of the bioreactor resulting in highly homogenous concentrations of the nutrients and metabolites in the whole system as confirmed by measurements from different sampling locations. A real-time biomass sensor using the dielectric properties of living cells was used to measure the cell density. The performances of the CellTank were studied in three perfusion runs. A very high cell density measured as 200 pF/cm (where 1 pF/cm is equivalent to 1 × 10(6)viable cells/mL) was achieved at a perfusion rate of 10 reactor volumes per day (RV/day) in the first run. In the second run, the effect of cell growth arrest by hypothermia at temperatures lowered gradually from 37 °C to 29 °C was studied during 13 days at cell densities above 100 pF/cm. Finally a production run was performed at high cell densities, where a temperature shift to 31 °C was applied at cell density 100 pF/cm during a production period of 14 days in minimized feeding conditions. The IgG concentrations were comparable in the matrix and in the harvest line in all the runs, indicating no retention of the product of interest. The cell specific productivity was comparable or higher than in Erlenmeyer flask batch culture. During the production run, the final harvested IgG production was 35 times higher in the CellTank compared to a repeated batch culture in the same vessel volume during the same time period. Copyright © 2015 The Authors. Published by Elsevier B.V. All

  11. Osteoblastic potency of bone marrow cells cultivated on functionalized biometals with cyclic RGD-peptide.

    Science.gov (United States)

    Jäger, M; Böge, C; Janissen, R; Rohrbeck, D; Hülsen, T; Lensing-Höhn, S; Krauspe, R; Herten, M

    2013-10-01

    The fixation of cementless endoprostheses requires excellent fixation at the bone implant interface. Although the surface structures of these implants are designed to promote osteoblastic differentiation, poor bone quality may prevent or delay osseointegration. There is evidence that RGD peptides known as recognition motifs for various integrins, promote cellular adhesion, influence cellular proliferation, and differentiation of local cells. In this study, five different metal surfaces were analyzed: Sandblasted (TiSa) and polished (TiPol) Ti6Al4V, porocoated (CCPor) and polished (CCPol) cobalt chrome and polished stainless steel (SS) were coated by ethanol amine and poly(ethylene glycol) to attach covalently RGD peptides. Human mesenchymal stromal cells of healthy donors were cultivated onto prior functionalized metal surfaces for 14 days without osteogenic stimulation. Cell proliferation and differentiation were quantitatively evaluated for native (I), NaOH pre-activated (II), NaOH pre-activated, and PEG-coated (III) as well as for RGD (IV) coated surfaces. The RGD immobilization efficiency was analyzed by epi-fluorescence spectroscopy, cell morphology was documented by light and scanning electron microscopy. The RGD-binding efficiency was TiSa > TiPol > SS > CCPor > CCPol. RGD coated surfaces showed the highest average cell proliferation on CCPol > SS > CCPor > TiSa ≥ TiPol, whereas cellular differentiation mostly correlated with the observed proliferation results, such as CCPol > TiSa > SS > CCPor > TiPol. Considering statistical analyses (significance level of α = 0.05), the RGD-coating of all biometals in comparison and in respect of their particular controls showed no significant improvement in cellular proliferation and osteoblastic differentiation. Copyright © 2013 Wiley Periodicals, Inc., a Wiley Company.

  12. Cytotoxic activity of ethanolic extracts of Eleutherococcus species cultivated in Poland on HL60 leukemia cell line

    Directory of Open Access Journals (Sweden)

    Zaluski Daniel

    2014-06-01

    Full Text Available The Eleutherococcus species including 40 species native to Asia are medicinal plants widely used in traditional medicine. Some of these species are cultivated at the botanical gardens in Europe. On the basis on our earlier studies it was concluded that the extracts of analyzed species act as antioxidants, inhibitors of MMPs, and cytotoxic against Jurkat 45 leukemia cell line. In this study, the anti-leukemic potential of roots and leaves from six Eleutherococcus species cultivated in Poland was determined. The in vitro cytotoxic activity towards human promyelotic leukemia cell line HL60 using trypan blue assay was evaluated. The induction of apoptosis in stimulated leukemia cells was determined by AnnexinV method. Morphological changes in treated cells were observed by microscopic investigations.

  13. Microvessel and mast cell densities in malignant laryngeal neoplasm

    Directory of Open Access Journals (Sweden)

    Balica Nicolae Constantin

    2014-01-01

    Full Text Available Laryngeal neoplasm contributes to 30-40% of carcinomas of the head and neck. Mast cells are normal connective tissue residents, well represented in the respiratory tract. Experimental evidence suggests that the growth of a tumor beyond a certain size requires angiogenesis, which may also permit metastasis. The aim of this study was to evaluate the correlation between mast cell density, microvascular density, histopathological type and histological grade. Our study included 38 laryngeal carcinomas as follows: adenoid cystic carcinoma (2 cases, malignant papilloma (2 cases and squamous cell carcinoma (34 cases. The combined technique of CD 34-alcian blue safranin (ABS was used to identify microvessel and mast cell density, which was quantified by the hot spot method. A significant correlation was found between both mast cell and microvascular density, and G1/G2 histological grade (p=0.002 and p=0.004, respectively. Squamous cell carcinoma was significantly correlated with mast cell density (p=0.003, but not with microvascular density (p=0.454.

  14. Long-term cultivation of human mammary carcionoma: proliferation and differential biochemical properties of the cultured cells.

    Science.gov (United States)

    Hakim, A A

    1979-01-01

    Human mammary carcinoma cell cultures proliferated from primary explants in Eagle's essential medium (MEM) supplemented with insulin, fetal calf serum (FCS) and/or human alpha-a1-antitrypsin. Human mammary carcinoma cells differed from normal mammary epithelial cells by the following catalytic activities: a. Thymidine uptake into the carcinoma cells was 6 to 10 fold greater, whereas thymidine conversion to CO2 was half to one fifth that of normal cells. b. The nucleolytic activity patterns of the mammary carcinoma cells preferred polycytydylic acid and double helical polynucleotides, whereas those of the normal mammary cells preferred polyuridylic acid and had no effect on double helical polynucleotides. c. The polymerase activity most evident in mammary carcinoma cells is a hybrid-dependent DNA polymerase which is guided by the ribo-strand of the template poly (rA) . poly(dT). In contrast the all-ribo template poly (rA) . poly(rU) showed little activity. d. There was slight or statistically non-significant difference between the amino acid composition of material cleaved from mammary carcinoma cells prepared from tumor tissues and from cells cultivated 10 months in vitro. e. There was no difference between the molar proportions of the carbohydrate components of the cell membrane from fresh tumor tissue and long term in vitro cultivated cells. f. The granules from long term in vitro cultured mammary carcinoma cells contained high collagenolytic, caseinolytic, fibrinolytic and esterolytic activities.

  15. Construction of an integrated high density simple sequence repeat linkage map in cultivated strawberry (Fragaria × ananassa) and its applicability.

    Science.gov (United States)

    Isobe, Sachiko N; Hirakawa, Hideki; Sato, Shusei; Maeda, Fumi; Ishikawa, Masami; Mori, Toshiki; Yamamoto, Yuko; Shirasawa, Kenta; Kimura, Mitsuhiro; Fukami, Masanobu; Hashizume, Fujio; Tsuji, Tomoko; Sasamoto, Shigemi; Kato, Midori; Nanri, Keiko; Tsuruoka, Hisano; Minami, Chiharu; Takahashi, Chika; Wada, Tsuyuko; Ono, Akiko; Kawashima, Kumiko; Nakazaki, Naomi; Kishida, Yoshie; Kohara, Mitsuyo; Nakayama, Shinobu; Yamada, Manabu; Fujishiro, Tsunakazu; Watanabe, Akiko; Tabata, Satoshi

    2013-02-01

    The cultivated strawberry (Fragaria × ananassa) is an octoploid (2n = 8x = 56) of the Rosaceae family whose genomic architecture is still controversial. Several recent studies support the AAA'A'BBB'B' model, but its complexity has hindered genetic and genomic analysis of this important crop. To overcome this difficulty and to assist genome-wide analysis of F. × ananassa, we constructed an integrated linkage map by organizing a total of 4474 of simple sequence repeat (SSR) markers collected from published Fragaria sequences, including 3746 SSR markers [Fragaria vesca expressed sequence tag (EST)-derived SSR markers] derived from F. vesca ESTs, 603 markers (F. × ananassa EST-derived SSR markers) from F. × ananassa ESTs, and 125 markers (F. × ananassa transcriptome-derived SSR markers) from F. × ananassa transcripts. Along with the previously published SSR markers, these markers were mapped onto five parent-specific linkage maps derived from three mapping populations, which were then assembled into an integrated linkage map. The constructed map consists of 1856 loci in 28 linkage groups (LGs) that total 2364.1 cM in length. Macrosynteny at the chromosome level was observed between the LGs of F. × ananassa and the genome of F. vesca. Variety distinction on 129 F. × ananassa lines was demonstrated using 45 selected SSR markers.

  16. Stability of high cell density brewery fermentations during serial repitching.

    Science.gov (United States)

    Verbelen, Pieter J; Dekoninck, Tinne M L; Van Mulders, Sebastiaan E; Saerens, Sofie M G; Delvaux, Filip; Delvaux, Freddy R

    2009-11-01

    The volumetric productivity of the beer fermentation process can be increased by using a higher pitching rate (i.e. higher inoculum size). However, the decreased yeast net growth observed in these high cell density brewery fermentations can adversely affect the physiological stability throughout subsequent yeast generations. Therefore, different O(2) conditions (wort aeration and yeast preoxygenation) were applied to high cell density fermentation and eight generations of fermentations were evaluated together with conventional fermentations. Freshly propagated high cell density populations adapted faster to the fermentative conditions than normal cell density populations. Preoxygenating the yeast was essential for the yeast physiological and beer flavor compound stability of high cell density fermentations during serial repitching. In contrast, the use of non-preoxygenated yeast resulted in inadequate growth which caused (1) insufficient yield of biomass to repitch all eight generations, (2) a 10% decrease in viability, (3) a moderate increase of yeast age, (4) and a dramatic increase of the unwanted flavor compounds acetaldehyde and total diacetyl during the sequence of fermentations. Therefore, to achieve sustainable high cell density fermentations throughout the economical valuable process of serial repitching, adequate yeast growth is essential.

  17. Cultivation of human neural progenitor cells in a 3-dimensional self-assembling peptide hydrogel.

    Science.gov (United States)

    Liedmann, Andrea; Rolfs, Arndt; Frech, Moritz J

    2012-01-11

    technique like fluorescence microscopes able to take z-stacks of the specimen. Furthermore this kind of analysis is extremely time consuming. Here we demonstrate a method to release cells from the 3D-scaffolds for the later analysis e.g. by flow cytometry. In this protocol human neural progenitor cells (hNPCs) of the ReNcell VM cell line (Millipore USA) were cultured and differentiated in 3D-scaffolds consisting of PuraMatrix (PM) or PuraMatrix supplemented with laminin (PML). In our hands a PM-concentration of 0.25% was optimal for the cultivation of the cells, however the concentration might be adapted to other cell types. The released cells can be used for e.g. immunocytochemical studies and subsequently analysed by flow cytometry. This speeds up the analysis and more over, the obtained data rest upon a wider base, improving the reliability of the data. Copyright © 2012 Journal of Visualized Experiments

  18. Cultivation of Human Bone-Like Tissue from Pluripotent Stem Cell-Derived Osteogenic Progenitors in Perfusion Bioreactors

    Science.gov (United States)

    de Peppo, Giuseppe Maria; Vunjak-Novakovic, Gordana; Marolt, Darja

    2014-01-01

    Human pluripotent stem cells represent an unlimited source of skeletal tissue progenitors for studies of bone biology, pathogenesis, and the development of new approaches for bone reconstruction and therapies. In order to construct in vitro models of bone tissue development and to grow functional, clinical-size bone substitutes for transplantation, cell cultivation in three-dimensional environments composed of porous osteoconductive scaffolds and dynamic culture systems—bioreactors—has been studied. Here, we describe a stepwise procedure for the induction of human embryonic and induced pluripotent stem cells (collectively termed PSCs) into mesenchymal-like progenitors, and their subsequent cultivation on decellularized bovine bone scaffolds in perfusion bioreactors, to support the development of viable, stable bone-like tissue in defined geometries. PMID:24281874

  19. Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities.

    Science.gov (United States)

    Mwachaka, Philip Maseghe; Saidi, Hassan; Odula, Paul Ochieng; Mandela, Pamela Idenya

    2015-01-01

    To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of the retina were taken and imported into FIJI software for analysis. Neural retinal cell densities of deprived eyes were reduced along with increasing period of deprivation. The percentage of reductions were 60.9% (P < 0.001), 41.6% (P = 0.003), and 18.9% (P = 0.326) for ganglion, inner nuclear, and outer nuclear cells, respectively. In non-deprived eyes, cell densities in contrast were increased by 116% (P < 0.001), 52% (P < 0.001) and 59.6% (P < 0.001) in ganglion, inner nuclear, and outer nuclear cells, respectively. In this rabbit model, monocular deprivation resulted in activity-dependent changes in cell densities of the neural retina in favour of the non-deprived eye along with reduced cell densities in the deprived eye.

  20. An inactivated yellow fever 17DD vaccine cultivated in Vero cell cultures.

    Science.gov (United States)

    Pereira, Renata C; Silva, Andrea N M R; Souza, Marta Cristina O; Silva, Marlon V; Neves, Patrícia P C C; Silva, Andrea A M V; Matos, Denise D C S; Herrera, Miguel A O; Yamamura, Anna M Y; Freire, Marcos S; Gaspar, Luciane P; Caride, Elena

    2015-08-20

    Yellow fever is an acute infectious disease caused by prototype virus of the genus Flavivirus. It is endemic in Africa and South America where it represents a serious public health problem causing epidemics of hemorrhagic fever with mortality rates ranging from 20% to 50%. There is no available antiviral therapy and vaccination is the primary method of disease control. Although the attenuated vaccines for yellow fever show safety and efficacy it became necessary to develop a new yellow fever vaccine due to the occurrence of rare serious adverse events, which include visceral and neurotropic diseases. The new inactivated vaccine should be safer and effective as the existing attenuated one. In the present study, the immunogenicity of an inactivated 17DD vaccine in C57BL/6 mice was evaluated. The yellow fever virus was produced by cultivation of Vero cells in bioreactors, inactivated with β-propiolactone, and adsorbed to aluminum hydroxide (alum). Mice were inoculated with inactivated 17DD vaccine containing alum adjuvant and followed by intracerebral challenge with 17DD virus. The results showed that animals receiving 3 doses of the inactivated vaccine (2 μg/dose) with alum adjuvant had neutralizing antibody titers above the cut-off of PRNT50 (Plaque Reduction Neutralization Test). In addition, animals immunized with inactivated vaccine showed survival rate of 100% after the challenge as well as animals immunized with commercial attenuated 17DD vaccine. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Expansion of Endothelial Progenitor Cells in High Density Dot Culture of Rat Bone Marrow Cells

    Science.gov (United States)

    Wang, Ling; Kretlow, James D.; Zhou, Guangdong; Cao, Yilin; Liu, Wei; Zhang, Wen Jie

    2014-01-01

    In vitro expansion of endothelial progenitor cells (EPCs) remains a challenge in stem cell research and its application. We hypothesize that high density culture is able to expand EPCs from bone marrow by mimicking cell-cell interactions of the bone marrow niche. To test the hypothesis, rat bone marrow cells were either cultured in high density (2×105 cells/cm2) by seeding total 9×105 cells into six high density dots or cultured in regular density (1.6×104 cells/cm2) with the same total number of cells. Flow cytometric analyses of the cells cultured for 15 days showed that high density cells exhibited smaller cell size and higher levels of marker expression related to EPCs when compared to regular density cultured cells. Functionally, these cells exhibited strong angiogenic potentials with better tubal formation in vitro and potent rescue of mouse ischemic limbs in vivo with their integration into neo-capillary structure. Global gene chip and ELISA analyses revealed up-regulated gene expression of adhesion molecules and enhanced protein release of pro-angiogenic growth factors in high density cultured cells. In summary, high density cell culture promotes expansion of bone marrow contained EPCs that are able to enhance tissue angiogenesis via paracrine growth factors and direct differentiation into endothelial cells. PMID:25254487

  2. A protocol for isolation and enriched monolayer cultivation of neural precursor cells from mouse dentate gyrus

    Directory of Open Access Journals (Sweden)

    Harish eBabu

    2011-07-01

    Full Text Available In vitro assays are valuable tools to study the characteristics of adult neural precursor cells under controlled conditions with a defined set of parameters. We here present a detailed protocol based on our previous original publication (Babu et al., Enriched monolayer precursor cell cultures from micro-dissected adult mouse dentate gyrus yield functional granule cell-like neurons, PLoS One 2007, 2:e388 to isolate neural precursor cells from the hippocampus of adult mice and maintain and propagate them as adherent monolayer cultures. The strategy is based on the use of Percoll density gradient centrifugation to enrich precursor cells from the micro-dissected dentate gyrus. Based on the expression of Nestin and Sox2, a culture-purity of more than 98% can be achieved. The cultures are expanded under serum-free conditions in Neurobasal A medium with addition of the mitogens EGF and FGF2 as well as the supplements Glutamax-1 and B27. Under differentiation conditions, the precursor cells reliably generate approximately 30% neurons with appropriate morphological, molecular and electrophysiological characteristics that might reflect granule cell properties as their in vivo counterpart. We also highlight potential modifications to the protocol.

  3. Human Plasma and Human Platelet-rich Plasma as a Substitute for Fetal Calf Serum during Long-term Cultivation of Mesenchymal Dental Pulp Stem Cells

    Directory of Open Access Journals (Sweden)

    Tereza Suchánková Kleplová

    2014-01-01

    Full Text Available Aims: Our aims were to isolate and cultivate mesenchymal dental pulp stem cells (DPSC in various media enriched with human blood components, and subsequently to investigate their basic biological properties. Methods: DPSC were cultivated in five different media based on α MEM containing different concentrations of human plasma (HP, platelet-rich plasma (PRP, or fetal calf serum (FCS. The DPSC biological properties were examined periodically. Results: We cultivated DPSC in the various cultivation media over 15 population doublings except for the medium supplemented with 10% HP. Our results showed that DPSC cultivated in medium supplemented with 10% PRP showed the shortest average population doubling time (DT (28.6 ± 4.6 hours, in contrast to DPSC cultivated in 10% HP which indicated the longest DT (156.2 ± 17.8 hours; hence this part of the experiment had been cancelled in the 6th passage. DPSC cultivated in media with 2% FCS+ITS (DT 47.3 ± 10.4 hours, 2% PRP (DT 40.1 ± 5.7 hours and 2% HP (DT 49.0 ± 15.2 hours showed almost the same proliferative activity. DPSC’s viability in the 9th passage was over 90% except for the DPSC cultivated in the 10% HP media. Conclusions: We proved that human blood components are suitable substitution for FCS in cultivation media for long-term DPSC cultivation.

  4. Human plasma and human platelet-rich plasma as a substitute for fetal calf serum during long-term cultivation of mesenchymal dental pulp stem cells.

    Science.gov (United States)

    Suchánková Kleplová, Tereza; Soukup, Tomáš; Řeháček, Vít; Suchánek, Jakub

    2014-01-01

    Our aims were to isolate and cultivate mesenchymal dental pulp stem cells (DPSC) in various media enriched with human blood components, and subsequently to investigate their basic biological properties. DPSC were cultivated in five different media based on α MEM containing different concentrations of human plasma (HP), platelet-rich plasma (PRP), or fetal calf serum (FCS). The DPSC biological properties were examined periodically. We cultivated DPSC in the various cultivation media over 15 population doublings except for the medium supplemented with 10% HP. Our results showed that DPSC cultivated in medium supplemented with 10% PRP showed the shortest average population doubling time (DT) (28.6 ± 4.6 hours), in contrast to DPSC cultivated in 10% HP which indicated the longest DT (156.2 ± 17.8 hours); hence this part of the experiment had been cancelled in the 6th passage. DPSC cultivated in media with 2% FCS+ITS (DT 47.3 ± 10.4 hours), 2% PRP (DT 40.1 ± 5.7 hours) and 2% HP (DT 49.0 ± 15.2 hours) showed almost the same proliferative activity. DPSC's viability in the 9th passage was over 90% except for the DPSC cultivated in the 10% HP media. We proved that human blood components are suitable substitution for FCS in cultivation media for long-term DPSC cultivation.

  5. The Effect of a p38 Mitogen-Activated Protein Kinase Inhibitor on Cellular Senescence of Cultivated Human Corneal Endothelial Cells.

    Science.gov (United States)

    Hongo, Akane; Okumura, Naoki; Nakahara, Makiko; Kay, EunDuck P; Koizumi, Noriko

    2017-07-01

    We have begun a clinical trial of a cell-based therapy for corneal endothelial dysfunction in Japan. The purpose of this study was to investigate the usefulness of a p38 MAPK inhibitor for prevention cellular senescence in cultivated human corneal endothelial cells (HCECs). HCECs of 10 donor corneas were divided and cultured with or without SB203580 (a p38 MAPK inhibitor). Cell density and morphology were evaluated by phase-contrast microscopy. Expression of function-related proteins was examined by immunofluorescent staining. Cellular senescence was evaluated by SA-β-gal staining and Western blotting for p16 and p21. Senescence-associated factors were evaluated by membrane blotting array, quantitative PCR, and ELISA. Phase-contrast microscopy showed a significantly higher cell density for HCECs cultured with SB203580 than without SB203580 (2623 ± 657 cells/mm2 and 1752 ± 628 cells/mm2, respectively). The HCECs cultured with SB203580 maintained a hexagonal morphology and expressed ZO-1, N-cadherin, and Na+/K+-ATPase in the plasma membrane, whereas the control HCECs showed an altered staining pattern for these marker proteins. HCECs cultured without SB203580 showed high positive SA-β-gal staining, a low nuclear/cytoplasm ratio, and expression of p16 and p21. IL-6, IL-8, CCL2, and CXCL1 were observed at high levels in low cell density HCECs cultured without SB203580. Activation of p38 MAPK signaling due to culture stress might be a causative factor that induces cellular senescence; therefore, the use of p38 MAPK inhibitor to counteract senescence may achieve sufficient numbers of HCECs for tissue engineering therapy for corneal endothelial dysfunction.

  6. Xeno-Free Cultivation of Mesenchymal Stem Cells From the Corneal Stroma.

    Science.gov (United States)

    Matthyssen, Steffi; Ní Dhubhghaill, Sorcha; Van Gerwen, Veerle; Zakaria, Nadia

    2017-05-01

    The human cornea has recently been described as a source of corneal stroma-derived mesenchymal stem cells (hMSCs). In vitro expansion of these cells involves basal medium supplemented with fetal bovine serum (FBS). As animal-derived serum can confer a risk of disease transmission and can be subject to considerable lot-to-lot variability, it does not comply with newer Good Manufacturing Practice (GMP)-required animal component-free culture protocols for clinical translation. This study investigated animal-free alternatives to FBS for cultivation of human corneal stromal MSCs. Proliferative capacity was studied for cultures supplemented with different concentrations (2.5%, 5%, and 10%) of FBS, human AB serum, human platelet lysate (HPL), and XerumFree. Unsupplemented basal medium was used as a control. The expression of specific hMSC markers (CD73+, CD90+, CD105+, CD19-, CD34-, CD79α-, CD11b-, CD14-, CD45-, and HLA-DR-) and trilineage differentiation (adipogenesis, osteogenesis, and chondrogenesis) were compared for the two outperforming supplements: 10% FBS and HPL. HPL is the only consistent non-xeno supplement where hMSC cultures show significantly higher proliferation than the 10% FBS-supplemented cultures. Both FBS- and HPL-supplemented hMSC cultures showed plastic adherence and trilineage differentiation, and no significant differences were found in the expression of the hMSC marker panel. No significant differences in stemness were detected between FBS and HPL cultures. We conclude that HPL is the best supplement for expansion of human corneal stromal MSCs. HPL significantly outperforms human AB serum, the chemically defined XerumFree, and even the gold standard, FBS. The xeno-free nature of HPL additionally confers preferred standing for use in GMP-regulated clinical trials using human corneal stromal MSCs.

  7. Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus

    Directory of Open Access Journals (Sweden)

    Daniel Reyes-Haro

    2016-01-01

    Full Text Available Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by anorexia is unknown. Thus, the aim of this study was to estimate GFAP+ cell density in the main regions of the hippocampus (CA1, CA2, CA3, and dentate gyrus in the DIA model. Our results showed that GFAP+ cell density was significantly reduced (~20% in all regions of the hippocampus, except in CA1. Interestingly, DIA significantly reduced the GFAP+ cells/nuclei ratio in CA2 (−23% and dentate gyrus (−48%. The reduction of GFAP+ cell density was in agreement with a lower expression of GFAP protein. Additionally, anorexia increased the expression of the intermediate filaments vimentin and nestin. Accordingly, anorexia increased the number of reactive astrocytes in CA2 and dentate gyrus more than twofold. We conclude that anorexia reduces the hippocampal GFAP+ cell density and increases vimentin and nestin expression.

  8. Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus.

    Science.gov (United States)

    Reyes-Haro, Daniel; Labrada-Moncada, Francisco Emmanuel; Varman, Durairaj Ragu; Krüger, Janina; Morales, Teresa; Miledi, Ricardo; Martínez-Torres, Ataúlfo

    2016-01-01

    Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA) is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by anorexia is unknown. Thus, the aim of this study was to estimate GFAP+ cell density in the main regions of the hippocampus (CA1, CA2, CA3, and dentate gyrus) in the DIA model. Our results showed that GFAP+ cell density was significantly reduced (~20%) in all regions of the hippocampus, except in CA1. Interestingly, DIA significantly reduced the GFAP+ cells/nuclei ratio in CA2 (-23%) and dentate gyrus (-48%). The reduction of GFAP+ cell density was in agreement with a lower expression of GFAP protein. Additionally, anorexia increased the expression of the intermediate filaments vimentin and nestin. Accordingly, anorexia increased the number of reactive astrocytes in CA2 and dentate gyrus more than twofold. We conclude that anorexia reduces the hippocampal GFAP+ cell density and increases vimentin and nestin expression.

  9. A protocol for isolation and enriched monolayer cultivation of neural precursor cells from mouse dentate gyrus.

    Science.gov (United States)

    Babu, Harish; Claasen, Jan-Hendrik; Kannan, Suresh; Rünker, Annette E; Palmer, Theo; Kempermann, Gerd

    2011-01-01

    In vitro assays are valuable tools to study the characteristics of adult neural precursor cells under controlled conditions with a defined set of parameters. We here present a detailed protocol based on our previous original publication (Babu et al., 2007) to isolate neural precursor cells from the hippocampus of adult mice and maintain and propagate them as adherent monolayer cultures. The strategy is based on the use of Percoll density gradient centrifugation to enrich precursor cells from the micro-dissected dentate gyrus. Based on the expression of Nestin and Sox2, a culture-purity of more than 98% can be achieved. The cultures are expanded under serum-free conditions in Neurobasal A medium with addition of the mitogens Epidermal growth factor and Fibroblast growth factor 2 as well as the supplements Glutamax-1 and B27. Under differentiation conditions, the precursor cells reliably generate approximately 30% neurons with appropriate morphological, molecular, and electrophysiological characteristics that might reflect granule cell properties as their in vivo counterpart. We also highlight potential modifications to the protocol.

  10. High volumetric power density, non-enzymatic, glucose fuel cells

    OpenAIRE

    Oncescu, Vlad; Erickson, David

    2013-01-01

    The development of new implantable medical devices has been limited in the past by slow advances in lithium battery technology. Non-enzymatic glucose fuel cells are promising replacement candidates for lithium batteries because of good long-term stability and adequate power density. The devices developed to date however use an ?oxygen depletion design? whereby the electrodes are stacked on top of each other leading to low volumetric power density and complicated fabrication protocols. Here we...

  11. Solid Oxide Electrolysis Cells: Degradation at High Current Densities

    DEFF Research Database (Denmark)

    Knibbe, Ruth; Traulsen, Marie Lund; Hauch, Anne

    2010-01-01

    The degradation of Ni/yttria-stabilized zirconia (YSZ)-based solid oxide electrolysis cells operated at high current densities was studied. The degradation was examined at 850°C, at current densities of −1.0, −1.5, and −2.0 A/cm2, with a 50:50 (H2O:H2) gas supplied to the Ni/YSZ hydrogen electrode...

  12. Waves in cell monolayer without proliferation: density determines cell velocity and wave celerity

    CERN Document Server

    Tlili, S; Li, B; Cardoso, O; Ladoux, B; Delanoë-Ayari, H; Graner, F

    2016-01-01

    Collective cell migration contributes to morphogenesis, wound healing or tumor metastasis. Culturing epithelial monolayers on a substrate is an in vitro configuration suitable to quantitatively characterize such tissue migration by measuring cell velocity, density and cell-substrate interaction force. Inhibiting cell division, we limit cell density increase and favor steady cell migration, while by using long narrow strips we stabilise the migrating front shape, so that we observe migration over a day or more. In the monolayer bulk, the cell velocity is a function of the cell density, namely it increases as a linear function of the cell radius. At least ten periods of propagating velocity waves are detected with a high signal-to-noise ratio, enabling for their quantitative spatio-temporal analysis. Cell density displays waves, in phase opposition with the velocity, as predicted by mass conservation; similarly, cell-substrate force appear to display small amplitude waves, in phase quadrature with respect to ve...

  13. C. albicans increases cell wall mannoprotein, but not mannan, in response to blood, serum and cultivation at physiological temperature.

    Science.gov (United States)

    Kruppa, Michael; Greene, Rachel R; Noss, Ilka; Lowman, Douglas W; Williams, David L

    2011-09-01

    The cell wall of Candida albicans is central to the yeasts ability to withstand osmotic challenge, to adhere to host cells, to interact with the innate immune system and ultimately to the virulence of the organism. Little is known about the effect of culture conditions on the cell wall structure and composition of C. albicans. We examined the effect of different media and culture temperatures on the molecular weight (Mw), polymer distribution and composition of cell wall mannan and mannoprotein complex. Strain SC5314 was inoculated from frozen stock onto yeast peptone dextrose (YPD), blood or 5% serum agar media at 30 or 37°C prior to mannan/mannoprotein extraction. Cultivation of the yeast in blood or serum at physiologic temperature resulted in an additive effect on Mw, however, cultivation media had the greatest impact on Mw. Mannan from a yeast grown on blood or serum at 30°C showed a 38.9 and 28.6% increase in Mw, when compared with mannan from YPD-grown yeast at 30°C. Mannan from the yeast pregrown on blood or serum at 37°C showed increased Mw (8.8 and 26.3%) when compared with YPD mannan at 37°C. The changes in Mw over the entire polymer distribution were due to an increase in the amount of mannoprotein (23.8-100%) and a decrease in cell wall mannan (5.7-17.3%). We conclude that C. albicans alters the composition of its cell wall, and thus its phenotype, in response to cultivation in blood, serum and/or physiologic temperature by increasing the amount of the mannoprotein and decreasing the amount of the mannan in the cell wall.

  14. Ultrahigh-density linkage map for cultivated cucumber (Cucumis sativus L. using a single-nucleotide polymorphism genotyping array.

    Directory of Open Access Journals (Sweden)

    Mor Rubinstein

    Full Text Available Genotyping arrays are tools for high-throughput genotyping, which is beneficial in constructing saturated genetic maps and therefore high-resolution mapping of complex traits. Since the report of the first cucumber genome draft, genetic maps have been constructed mainly based on simple-sequence repeats (SSRs or on combinations of SSRs and sequence-related amplified polymorphism (SRAP. In this study, we developed the first cucumber genotyping array consisting of 32,864 single-nucleotide polymorphisms (SNPs. These markers cover the cucumber genome with a median interval of ~2 Kb and have expected genotype calls in parents/F1 hybridizations as a training set. The training set was validated with Fluidigm technology and showed 96% concordance with the genotype calls in the parents/F1 hybridizations. Application of the genotyping array was illustrated by constructing a 598.7 cM genetic map based on a '9930' × 'Gy14' recombinant inbred line (RIL population comprised of 11,156 SNPs. Marker collinearity between the genetic map and reference genomes of the two parents was estimated at R2 = 0.97. We also used the array-derived genetic map to investigate chromosomal rearrangements, regional recombination rate, and specific regions with segregation distortions. Finally, 82% of the linkage-map bins were polymorphic in other cucumber variants, suggesting that the array can be applied for genotyping in other lines. The genotyping array presented here, together with the genotype calls of the parents/F1 hybridizations as a training set, should be a powerful tool in future studies with high-throughput cucumber genotyping. An ultrahigh-density linkage map constructed by this genotyping array on RIL population may be invaluable for assembly improvement, and for mapping important cucumber QTLs.

  15. High Energy Density Regenerative Fuel Cell Systems for Terrestrial Applications

    Science.gov (United States)

    Burke, Kenneth A.

    1999-01-01

    Regenerative Fuel Cell System (RFCS) technology for energy storage has been a NASA power system concept for many years. Compared to battery-based energy storage systems, RFCS has received relatively little attention or resources for development because the energy density and electrical efficiency were not sufficiently attractive relative to advanced battery systems. Even today, RFCS remains at a very low technology readiness level (TRL of about 2 indicating feasibility has been demonstrated). Commercial development of the Proton Exchange Membrane (PEM) fuel cells for automobiles and other terrestrial applications and improvements in lightweight pressure vessel design to reduce weight and improve performance make possible a high energy density RFCS energy storage system. The results from this study of a lightweight RFCS energy storage system for a remotely piloted, solar-powered, high altitude aircraft indicate an energy density up to 790 w-h/kg with electrical efficiency of 53.4% is attainable. Such an energy storage system would allow a solar-powered aircraft to carry hundreds of kilograms of payload and remain in flight indefinitely for use in atmospheric research, earth observation, resource mapping. and telecommunications. Future developments in the areas of hydrogen and oxygen storage, pressure vessel design, higher temperature and higher- pressure fuel cell operation, unitized regenerative fuel cells, and commercial development of fuel cell technology will improve both the energy density and electrical efficiency of the RFCS.

  16. Evolvement of cell-substrate interaction over time for cells cultivated on a 3-aminopropyltriethoxysilane ({gamma}-APTES) modified silicon dioxide (SiO{sub 2}) surface

    Energy Technology Data Exchange (ETDEWEB)

    Hsu, Chung-Ping [Division of Thoracic Surgery, Department of Surgery, Taichung Veterans General Hospital, Taichung, 40705, Taiwan, ROC (China); School of Medicine, National Yang Ming University, Taipei, 11221, Taiwan, ROC (China); Hsu, Po-Yen [Department of Electrical Engineering, National Chi Nan University, Puli, Nantou, 54561, Taiwan, ROC (China); Wu, You-Lin, E-mail: ylwu@ncnu.edu.tw [Department of Electrical Engineering, National Chi Nan University, Puli, Nantou, 54561, Taiwan, ROC (China); Hsu, Wan-Yun [Comprehensive Cancer Center, Taichung Veterans General Hospital, Taichung, 40705, Taiwan, ROC (China); Lin, Jing-Jenn [Department of Applied Materials and Optoelectronic Engineering, National Chi Nan University, Puli, Nantou, 54561, Taiwan, ROC (China)

    2012-09-01

    Highlights: Black-Right-Pointing-Pointer Cell-substrate interaction of single cells was observed directly from the post-cell-removal imprint left on {gamma}-APTES soft substrate surface. Black-Right-Pointing-Pointer The time evolvement of the cell-substrate interaction can be obtained by cultivating cells on the {gamma}-APTES surface for different periods of time. Black-Right-Pointing-Pointer The cell-substrate interaction property can be found from the post-cell-removal surface morphology profiles determined by AFM. Black-Right-Pointing-Pointer It was found that the cancer cells tend to form deeper trenches along the circumference of the imprints, while the normal cells do not. - Abstract: Since cell-substrate interaction is directly related to the traction force of the cell, the cell property can be judged from the imprint it leaves on the soft substrate surface onto which the cell is cultured. In this letter, the evolvement of the cell-substrate interaction over time was observed by cultivating cells on a 3-aminopropyltriethoxysilane ({gamma}-APTES) modified silicon dioxide (SiO{sub 2}) surface for different periods of time. The cell-substrate interaction property as a function of time can then be found from the post-cell-removal surface morphology profiles determined by atomic force microscopy (AFM). Different surface morphology profiles were found between normal cells and cancer cells. It was found that the cancer cells tend to form deeper trenches along the circumference of the imprints, while the normal cells do not. In addition, our results indicated that normal cells involve cell-substrate interaction mechanisms that are different from those for cancer cells.

  17. Measuring density and compressibility of white blood cells and prostate cancer cells by microchannel acoustophoresis

    DEFF Research Database (Denmark)

    Barnkob, Rune; Augustsson, Per; Magnusson, Cecilia

    2011-01-01

    to determine the density and compressibility of individual cells enables the prediction and alteration of the separation outcome for a given cell mixture. We apply the method on white blood cells (WBCs) and DU145 prostate cancer cells (DUCs) aiming to improve isolation of circulating tumor cells from blood......, an emerging tool in the monitoring and characterizing of metastatic cancer....

  18. Any Defining Role of Mast Cell or Mast Cell Density in Oral ...

    African Journals Online (AJOL)

    Mast Cell Density in. Oral Squamous Cell. Carcinoma? Dear Sir,. I read an article by Zaidi et al. titled to “A study on assessment of mast cell (MCs) in oral squamous cell carcinoma (OSCC)” with great interest.[1] We are concerned about ... ovarian cancer.[11] Furthermore, chymase induces apoptosis in different types of cells ...

  19. Response of Escherichia coli to nutrient availability during cultivation at single cell level

    DEFF Research Database (Denmark)

    Han, Shanshan

    -limited Saccharomyces cerevisiae and Escherichia coli chemostat cultures. The physiology of S. cerevisiae and E. coli at low growth rates deserves more attention as slow-growing cells were more robust towards stress than fast-growing cells. A high GFP expression and more dynamic changes were observed in slow growing......Industrial fermentation processes comprise the potential sources of heterogeneity since the microbial cells exhibit an intrinsic cell-to-cell variability that is both affected by and influence the fermentation process itself (reactor conditions and medium composition) over time. However, population...... promoters were related to the cultivation mode with different glucose limitation. The sudden glucose relief elevated the ribosomal RNA synthesis as well as its transcriptional activator Fis. Heterogeneities induced by scale-down may affect protein leakage into the broth medium because of the increased...

  20. High volumetric power density, non-enzymatic, glucose fuel cells

    Science.gov (United States)

    Oncescu, Vlad; Erickson, David

    2013-01-01

    The development of new implantable medical devices has been limited in the past by slow advances in lithium battery technology. Non-enzymatic glucose fuel cells are promising replacement candidates for lithium batteries because of good long-term stability and adequate power density. The devices developed to date however use an “oxygen depletion design” whereby the electrodes are stacked on top of each other leading to low volumetric power density and complicated fabrication protocols. Here we have developed a novel single-layer fuel cell with good performance (2 μW cm−2) and stability that can be integrated directly as a coating layer on large implantable devices, or stacked to obtain a high volumetric power density (over 16 μW cm−3). This represents the first demonstration of a low volume non-enzymatic fuel cell stack with high power density, greatly increasing the range of applications for non-enzymatic glucose fuel cells. PMID:23390576

  1. Multifrequency permittivity measurements enable on-line monitoring of changes in intracellular conductivity due to nutrient limitations during batch cultivations of CHO cells.

    Science.gov (United States)

    Ansorge, Sven; Esteban, Geoffrey; Schmid, Georg

    2010-01-01

    Lab and pilot scale batch cultivations of a CHO K1/dhfr(-) host cell line were conducted to evaluate on-line multifrequency permittivity measurements as a process monitoring tool. The beta-dispersion parameters such as the characteristic frequency (f(C)) and the permittivity increment (Deltaepsilon(max)) were calculated on-line from the permittivity spectra. The dual-frequency permittivity signal correlated well with the off-line measured biovolume and the viable cell density. A significant drop in permittivity was monitored at the transition from exponential growth to a phase with reduced growth rate. Although not reflected in off-line biovolume measurements, this decrease coincided with a drop in OUR and was probably caused by the depletion of glutamine and a metabolic shift occurring at the same time. Sudden changes in cell density, cell size, viability, capacitance per membrane area (C(M)), and effects caused by medium conductivity (sigma(m)) could be excluded as reasons for the decrease in permittivity. After analysis of the process data, a drop in f(C) as a result of a fall in intracellular conductivity (sigma(i)) was identified as responsible for the observed changes in the dual-frequency permittivity signal. It is hypothesized that the beta-dispersion parameter f(C) is indicative of changes in nutrient availability that have an impact on intracellular conductivity sigma(i). On-line permittivity measurements consequently not only reflect the biovolume but also the physiological state of mammalian cell cultures. These findings should pave the way for a better understanding of the intracellular state of cells and render permittivity measurements an important tool in process development and control.

  2. Regional alterations in purkinje cell density in patients with autism.

    Directory of Open Access Journals (Sweden)

    Jerry Skefos

    Full Text Available Neuropathological studies, using a variety of techniques, have reported a decrease in Purkinje cell (PC density in the cerebellum in autism. We have used a systematic sampling technique that significantly reduces experimenter bias and variance to estimate PC densities in the postmortem brains of eight clinically well-documented individuals with autism, and eight age- and gender-matched controls. Four cerebellar regions were analyzed: a sensorimotor area comprised of hemispheric lobules IV-VI, crus I & II of the posterior lobe, and lobule X of the flocculonodular lobe. Overall PC density was thus estimated using data from all three cerebellar lobes and was found to be lower in the cases with autism as compared to controls, an effect that was most prominent in crus I and II (p<0.05. Lobule X demonstrated a trend towards lower PC density in only the males with autism (p = 0.05. Brain weight, a correlate of tissue volume, was found to significantly contribute to the lower lobule X PC density observed in males with autism, but not to the finding of lower PC density in crus I & II. Therefore, lower crus I & II PC density in autism is more likely due to a lower number of PCs. The PC density in lobule X was found to correlate with the ADI-R measure of the patient's use of social eye contact (R² = -0.75, p = 0.012. These findings support the hypothesis that abnormal PC density may contribute to selected clinical features of the autism phenotype.

  3. Effect of cell density on adipogenic differentiation of mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Hongxu [Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577 (Japan); Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); Guo, Likun [Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); National Engineering Research Center for Biomaterials, Sichuan University, 29 Wangjiang Road, Chengdu 610064 (China); Wozniak, Michal J. [Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); Kawazoe, Naoki [Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); International Center for Materials Nanoarchitectonics (MANA), National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); Tateishi, Tetsuya [Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); Zhang, Xingdong [National Engineering Research Center for Biomaterials, Sichuan University, 29 Wangjiang Road, Chengdu 610064 (China); Chen, Guoping, E-mail: Guoping.CHEN@nims.go.jp [Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8577 (Japan); Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan); International Center for Materials Nanoarchitectonics (MANA), National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 (Japan)

    2009-04-10

    The effect of cell density on the adipogenic differentiation of human bone marrow-derived mesenchymal stem cells (MSCs) was investigated by using a patterning technique to induce the formation of a cell density gradient on a micropatterned surface. The adipogenic differentiation of MSCs at a density gradient from 5 x 10{sup 3} to 3 x 10{sup 4} cells/cm{sup 2} was examined. Lipid vacuoles were observed at all cell densities after 1-3 weeks of culture in adipogenic differentiation medium although the lipid vacuoles were scarce at the low cell density and abundant at the high cell density. Real-time RT-PCR analysis showed that adipogenesis marker genes encoding peroxisome proliferator-activated receptor {gamma}2 (PPAR{gamma}2), lipoprotein lipase (LPL), and fatty acid binding protein-4 (FABP4) were detected in the MSCs cultured at all cell densities. The results suggest that there was no apparent effect of cell density on the adipogenic differentiation of human MSCs.

  4. Corneal endothelial cell density and morphology in normal Iranian eyes

    Directory of Open Access Journals (Sweden)

    Fallah Mohammad

    2006-03-01

    Full Text Available Abstract Background We describe corneal endothelial cell density and morphology in normal Iranian eyes and compare endothelial cell characteristics in the Iranian population with data available in the literature for American and Indian populations. Methods Specular microscopy was performed in 525 eyes of normal Iranian people aged 20 to 85 years old. The studied parameters including mean endothelial cell density (MCD, mean cell area (MCA and coefficient of variation (CV in cell area were analyzed in all of the 525 eyes. Results MCD was 1961 ± 457 cell/mm2 and MCA was 537.0 ± 137.4 μm2. There was no statistically significant difference in MCD, MCA and CV between genders (Student t-test, P = 0.85, P = 0.97 and P = 0.15 respectively. There was a statistically significant decrease in MCD with age (P r = -0.64. The rate of cell loss was 0.6% per year. There was also a statistically significant increase in MCA (P r = 0.56 and CV (P r = 0.30 from 20 to 85 years of age. Conclusion The first normative data for the endothelium of Iranian eyes seems to confirm that there are no differences in MCD, MCA and CV between genders. Nevertheless, the values obtained in Iranian eyes seem to be different to those reported by the literature in Indian and American populations.

  5. [Comparison of the indirect immunofluorescence assay performance of Bartonella henselae antigens obtained by co-cultivation in Vero and HeLa cells].

    Science.gov (United States)

    Ergin, Cağrı; Akkaya, Yüksel; Kiriş Satılmış, Ozgün; Yılmaz, Cansev

    2011-07-01

    The laboratory diagnosis of Bartonella henselae infection is mainly based on serological testing by indirect immunofluorescence assay (IFA). Cell line co-cultivation with B.henselae and agar derivated antigens are the two major procedures used for evaluation of anti-Bartonella antibodies. Vero and Hep-2 cell lines are the most commonly used media for co-cultivation both in-house and commercial diagnostic kits production. However, HeLa cells which are easily supplied and grown, also can easily be infected by B.henselae. The aim of this study was to compare the performances of antigens obtained by co-cultivation of B.henselae ATCC 49882 (Houston-1) in Vero and HeLa Cells in IFA serology. Out of 381 sera samples, 127 (33.3%) were found positive and 195 (51.2%) were found negative by IFA performed by both cell line co-cultivations. The total agreement between the methods were found as 84.5% (322/381), and Cohen kappa value was calculated as 0.68 (strong, coherent). As a result, He-La cells were found to be useful for the preparation of B.henselae antigens to be used in IFA for the serologic diagnosis of B.henselae infections. However different genotype strains and cross reactions with other infectious agents should be investigated by further studies before routine applications of HeLa cell co-cultivations procedure is established.

  6. Ocular Surface Reconstruction with Cultivated Limbal Epithelial Cells in Limbal Stem Cell Deficiency: One-year Follow-up Results

    Directory of Open Access Journals (Sweden)

    İsmet Durak

    2012-05-01

    Full Text Available Pur po se: To evaluate the 1-year follow-up results of cultivated limbal epithelial cell (CLEC transplantation in unilateral limbal stem cell deficiency (LSCD. Ma te ri al and Met hod: One-year follow-up results of five unilateral LSCD patients who had undergone CLEC transplantation were evaluated. Parameters for this evaluation were: fluorescein staining of ocular surface, corneal vascularization and status of epithelium with slit lamp, and visual acuity. 1.5-mm limbal biopsy was performed from the superior limbus of the healthy eyes, broke into two equal pieces, expanded on human amniotic membrane (hAM and inserts for 14 days until getting 20 mm in size. CLECs on hAMs were used directly, and cells on inserts were usedafter detachment procedure. The symblepharon and pannus tissues were removed, superficial keratectomy was performed. CLEC on hAMs were transplanted with the epithelial side up onto the bare corneal stroma, sutured to the conjunctiva with 10-0 nylon sutures. Free CLEC layer from insert was placed on hAM as a second layer, additional hAM was used as a protective layer all over other tissues. Re sults: Median age was 44.4 years (14-71. The etiology was chemical burn in all patients. Median duration of symptoms was 10 years (2-18, median follow-up period was 12.6 (12-12.5 months. Preoperative best corrected visual acuities (BCVA were light perception in three patients, counting fingers at 50 cm in one patient and 3/10 in one patient. Visions were improved in all patients. Postoperative BCVA 12 months after the surgery were between counting fingers at 3 meters to 6/10. There was a temporary hemorrhage between the two layers of hAMs in one patient at the early postoperative period. Peripheral corneal vascularization has occurred in three patients, in patient corneal vascularization has reached to the paracentral area. Dis cus si on: CLEC transplantation is an efficient treatment option for unilateral LSCD in mid-long term. (Turk J

  7. Nanofiber density determines endothelial cell behavior on hydrogel matrix

    Energy Technology Data Exchange (ETDEWEB)

    Berti, Fernanda V., E-mail: fernanda@intelab.ufsc.br [Department of Chemical and Food Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Rambo, Carlos R. [Department of Electrical Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Dias, Paulo F. [Department of Cell Biology, Embryology and Genetics, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil); Porto, Luismar M. [Department of Chemical and Food Engineering, Federal University of Santa Catarina, 88040-900 Florianópolis, SC (Brazil)

    2013-12-01

    When cultured under static conditions, bacterial cellulose pellicles, by the nature of the polymer synthesis that involves molecular oxygen, are characterized by two distinct surface sides. The upper surface is denser in fibers (entangled) than the lower surface that shows greater surface porosity. Human umbilical vein endothelial cells (HUVECs) were used to exploit how the microarchitecture (i.e., surface porosity, fiber network structure, surface topology, and fiber density) of bacterial cellulose pellicle surfaces influence cell–biomaterial interaction and therefore cell behavior. Adhesion, cell ingrowth, proliferation, viability and cell death mechanisms were evaluated on the two pellicle surface sides. Cell behavior, including secondary necrosis, is influenced only by the microarchitecture of the surface, since the biomaterial is extremely pure (constituted of cellulose and water only). Cell–cellulose fiber interaction is the determinant signal in the cell–biomaterial responses, isolated from other frequently present interferences such as protein and other chemical traces usually present in cell culture matrices. Our results suggest that microarchitecture of hydrogel materials might determine the performance of biomedical products, such as bacterial cellulose tissue engineering constructs (BCTECs). - Highlights: • Topography of BC pellicle is relevant to determine endothelial cells' fate. • Cell–biomaterial response is affected by the topography of BC-pellicle surface. • Endothelial cells exhibit different behavior depending on the BC topography. • Apoptosis and necrosis of endothelial cells were affected by the BC topography.

  8. Effect of Monocular Deprivation on Rabbit Neural Retinal Cell Densities

    OpenAIRE

    Philip Maseghe Mwachaka; Hassan Saidi; Paul Ochieng Odula; Pamela Idenya Mandela

    2015-01-01

    Purpose: To describe the effect of monocular deprivation on densities of neural retinal cells in rabbits. Methods: Thirty rabbits, comprised of 18 subject and 12 control animals, were included and monocular deprivation was achieved through unilateral lid suturing in all subject animals. The rabbits were observed for three weeks. At the end of each week, 6 experimental and 3 control animals were euthanized, their retinas was harvested and processed for light microscopy. Photomicrographs of ...

  9. Uncovering the cultivable microbial diversity of costa rican beetles and its ability to break down plant cell wall components.

    Directory of Open Access Journals (Sweden)

    Gabriel Vargas-Asensio

    Full Text Available Coleopterans are the most diverse insect order described to date. These organisms have acquired an array of survival mechanisms through their evolution, including highly efficient digestive systems. Therefore, the coleopteran intestinal microbiota constitutes an important source of novel plant cell wall-degrading enzymes with potential biotechnological applications. We isolated and described the cultivable fungi, actinomycetes and aerobic eubacteria associated with the gut of larvae and adults from six different beetle families colonizing decomposing logs in protected Costa Rican ecosystems. We obtained 611 isolates and performed phylogenetic analyses using the ITS region (fungi and 16S rDNA (bacteria. The majority of fungal isolates belonged to the order Hypocreales (26% of 169 total, while the majority of actinomycetes belonged to the genus Streptomyces (86% of 241 total. Finally, we isolated 201 bacteria spanning 19 different families belonging into four phyla: Firmicutes, α, β and γ-proteobacteria. Subsequently, we focused on microbes isolated from Passalid beetles to test their ability to degrade plant cell wall polymers. Highest scores in these assays were achieved by a fungal isolate (Anthostomella sp., two Streptomyces and one Bacillus bacterial isolates. Our study demonstrates that Costa Rican beetles harbor several types of cultivable microbes, some of which may be involved in symbiotic relationships that enable the insect to digest complex polymers such as lignocellulose.

  10. Uncovering the Cultivable Microbial Diversity of Costa Rican Beetles and Its Ability to Break Down Plant Cell Wall Components

    Science.gov (United States)

    Vargas-Asensio, Gabriel; Pinto-Tomas, Adrian; Rivera, Beatriz; Hernandez, Myriam; Hernandez, Carlos; Soto-Montero, Silvia; Murillo, Catalina; Sherman, David H.; Tamayo-Castillo, Giselle

    2014-01-01

    Coleopterans are the most diverse insect order described to date. These organisms have acquired an array of survival mechanisms through their evolution, including highly efficient digestive systems. Therefore, the coleopteran intestinal microbiota constitutes an important source of novel plant cell wall-degrading enzymes with potential biotechnological applications. We isolated and described the cultivable fungi, actinomycetes and aerobic eubacteria associated with the gut of larvae and adults from six different beetle families colonizing decomposing logs in protected Costa Rican ecosystems. We obtained 611 isolates and performed phylogenetic analyses using the ITS region (fungi) and 16S rDNA (bacteria). The majority of fungal isolates belonged to the order Hypocreales (26% of 169 total), while the majority of actinomycetes belonged to the genus Streptomyces (86% of 241 total). Finally, we isolated 201 bacteria spanning 19 different families belonging into four phyla: Firmicutes, α, β and γ-proteobacteria. Subsequently, we focused on microbes isolated from Passalid beetles to test their ability to degrade plant cell wall polymers. Highest scores in these assays were achieved by a fungal isolate (Anthostomella sp.), two Streptomyces and one Bacillus bacterial isolates. Our study demonstrates that Costa Rican beetles harbor several types of cultivable microbes, some of which may be involved in symbiotic relationships that enable the insect to digest complex polymers such as lignocellulose. PMID:25411842

  11. Peptone-yeast autolysate-fetal bovine serum 10, a simple, inexpensive liquid medium for cultivation of Leishmania spp.

    OpenAIRE

    Palomino, J. C.

    1982-01-01

    A simple liquid medium for the cultivation of Leishmania parasites is described. Leishmania brasiliensis and Leishmania peruviana cultured in this medium reached cell densities greater than 10(7) promastigotes per ml within 7 days. This medium compares very favorably with the more complex media used to cultivate Leishmania spp. and other hemoflagellates.

  12. Co-cultivation of fungal and microalgal cells as an efficient system for harvesting microalgal cells, lipid production and wastewater treatment.

    Directory of Open Access Journals (Sweden)

    Digby Wrede

    Full Text Available The challenges which the large scale microalgal industry is facing are associated with the high cost of key operations such as harvesting, nutrient supply and oil extraction. The high-energy input for harvesting makes current commercial microalgal biodiesel production economically unfeasible and can account for up to 50% of the total cost of biofuel production. Co-cultivation of fungal and microalgal cells is getting increasing attention because of high efficiency of bio-flocculation of microalgal cells with no requirement for added chemicals and low energy inputs. Moreover, some fungal and microalgal strains are well known for their exceptional ability to purify wastewater, generating biomass that represents a renewable and sustainable feedstock for biofuel production. We have screened the flocculation efficiency of the filamentous fungus A. fumigatus against 11 microalgae representing freshwater, marine, small (5 µm, large (over 300 µm, heterotrophic, photoautotrophic, motile and non-motile strains. Some of the strains are commercially used for biofuel production. Lipid production and composition were analysed in fungal-algal pellets grown on media containing alternative carbon, nitrogen and phosphorus sources contained in wheat straw and swine wastewater, respectively. Co-cultivation of algae and A. fumigatus cells showed additive and synergistic effects on biomass production, lipid yield and wastewater bioremediation efficiency. Analysis of fungal-algal pellet's fatty acids composition suggested that it can be tailored and optimised through co-cultivating different algae and fungi without the need for genetic modification.

  13. Co-Cultivation of Fungal and Microalgal Cells as an Efficient System for Harvesting Microalgal Cells, Lipid Production and Wastewater Treatment

    Science.gov (United States)

    Wrede, Digby; Taha, Mohamed; Miranda, Ana F.; Kadali, Krishna; Stevenson, Trevor; Ball, Andrew S.; Mouradov, Aidyn

    2014-01-01

    The challenges which the large scale microalgal industry is facing are associated with the high cost of key operations such as harvesting, nutrient supply and oil extraction. The high-energy input for harvesting makes current commercial microalgal biodiesel production economically unfeasible and can account for up to 50% of the total cost of biofuel production. Co-cultivation of fungal and microalgal cells is getting increasing attention because of high efficiency of bio-flocculation of microalgal cells with no requirement for added chemicals and low energy inputs. Moreover, some fungal and microalgal strains are well known for their exceptional ability to purify wastewater, generating biomass that represents a renewable and sustainable feedstock for biofuel production. We have screened the flocculation efficiency of the filamentous fungus A. fumigatus against 11 microalgae representing freshwater, marine, small (5 µm), large (over 300 µm), heterotrophic, photoautotrophic, motile and non-motile strains. Some of the strains are commercially used for biofuel production. Lipid production and composition were analysed in fungal-algal pellets grown on media containing alternative carbon, nitrogen and phosphorus sources contained in wheat straw and swine wastewater, respectively. Co-cultivation of algae and A. fumigatus cells showed additive and synergistic effects on biomass production, lipid yield and wastewater bioremediation efficiency. Analysis of fungal-algal pellet's fatty acids composition suggested that it can be tailored and optimised through co-cultivating different algae and fungi without the need for genetic modification. PMID:25419574

  14. T cells, mast cells and microvascular density in diffuse large B cell lymphoma.

    Science.gov (United States)

    Marinaccio, Christian; Ingravallo, Giuseppe; Gaudio, Francesco; Perrone, Tommasina; Ruggieri, Simona; Opinto, Giuseppina; Nico, Beatrice; Maiorano, Eugenio; Specchia, Giorgina; Ribatti, Domenico

    2016-08-01

    Diffuse large B cell lymphoma (DLBCL) is recognized as the most common form of non-Hodgkin lymphoma (NHL), accounting for about 40 % of all cases of NHL. Among the cellular components of the tumor inflammatory infiltrate, T cells and mast cells have been demonstrated to be correlated with tumor angiogenesis. In this report, we have investigated CD3 and tryptase expression and their relationship with microvascular density (MVD) in DLBCL patients. Moreover, we determined the significance of CD3 expression in bulky and non-bulky disease. CD3 expression was significantly lower in bulky disease patients when compared to non-bulky ones. CD3 showed a positive correlation with tryptase and MVD, while multiple regression analysis efficaciously predicted MVD depending on CD3 and tryptase as predictors, supporting a complex interplay between these cells in sustaining tumor angiogenesis in DLBCL patients.

  15. Does corneal hysteresis correlate with endothelial cell density?

    Science.gov (United States)

    Akova-Budak, Berna; Kıvanç, Sertaç Argun

    2015-05-21

    Our aim was to determine if there is a correlation between corneal biomechanical properties, endothelial cell count, and corneal pachymetry in healthy corneas. Ninety-two eyes of all subjects underwent complete ocular examination, including intraocular pressure measurement by Goldmann applanation tonometer, objective refraction, and slit-lamp biomicroscopy. Topographic measurements and corneal pachymetry were performed using a Scheimpflug-based (Pentacam, Oculus, Germany) corneal topographer. Corneal hysteresis (CH) and corneal resistance factor (CRF) were measured with an Ocular Response Analyzer (ORA, Reichert Ophthalmic Instruments, Buffalo, NY). Endothelial cell count measurement was done using a specular microscope (CellChek, Konan, USA). Right eye values of the subjects were taken for the study. The mean CH was 11.5±1.7 mmHg and the mean CRF was 11.2±1.4 mmHg. Mean intraocular pressure was 15.3±2.3 mmHg. The mean endothelial cell count was 2754±205 cells/mm2. No correlation was found between biomechanical properties of cornea and endothelial cell count. There was a significant positive correlation between CH, CRF, and corneal thickness (pcorneal biomechanical properties significantly correlated with corneal thickness. We found no correlation between CH and CRF with the endothelial cell density in normal subjects.

  16. [Comparative analysis of a new human cell line 4BL karyotype at long-term cultivation. Ploidy of chromosomal set].

    Science.gov (United States)

    Akopian, H R; Huleiuk, N L; Kushniruk, V O; Mykytenko, D O; Iatsyshyna, A P; Lukash, L L

    2013-01-01

    Long-term cultivation of human cells, including stem cells, can lead to substantial transformation of the karyotype and occurrence of genetic instability. The aim of this research was a comparative cytogenetic study of the karyotype of a new human stem cell line 4BL at 160 and 205 passages. The absence of 10 and 13 pairs of chromosomes and the monosomy of chromosomes 4, 8, 10, 11, 13, 15, 17, 21, X were observed; also six regular marker chromosomes were detected. Chromosomes 1, 15 and 21 are involved in translocations t(l;11), t(5;15), t(12; 15), t(16;21). Modal class of the karyotype is within 41-43 chromosomes at both 160 and 205 passages. The frequency of polyploid cells have been increased from 2.8% at 160 passage up to 36% at 205 passage. Cells with a near-haploid karyotype were not detected at 205 passage (in contrast to 24.6% at 160 passages) and a decline of the level of premature separation of chromatids was observed. We assume stabilization of karyotype of the cell line 4BL at 205 passage and consider that further research is needed to predict the direction of karyotypic evolution of these cells in vitro.

  17. Immunohistochemical analysis of mast cell infiltrates and microvessel density in oral squamous cell carcinoma.

    Science.gov (United States)

    Pyziak, L; Stasikowska-Kanicka, O; Danilewicz, M; Wągrowska-Danilewicz, M

    2013-12-01

    The aim of the study was to evaluate mast cell concentration and microvessel density in perilesional and intralesional regions of oral squamous cell carcinoma (OSCC) and furthermore to assess the possible relationship between the above-mentioned parameters. Paraffin-embedded specimens from 47 cases of OSCC and 12 cases of normal mucosa were investigated immunohistochemically with anti-CD-31 antibody to stain microvessels and anti-tryptase antibody to visualize mast cells. The degree of vascularization and mast cell infiltration was measured with an image analysis system. The study revealed considerably increased microvessel density and mast cell abundance in intralesional and perilesional regions of OSCCs in comparison to normal mucosa. There was a significant positive correlation between microvessel density and mast cell concentration in both localizations of OSCCs (p therapeutic significance which require further research.

  18. Cell density modulates intracellular mass transport in neural networks.

    Science.gov (United States)

    Cintora, Patricia; Arikkath, Jyothi; Kandel, Mikhail; Popescu, Gabriel; Best-Popescu, Catherine

    2017-05-01

    In order to fully understand brain connectivity and elucidate the mechanisms involved in central nervous system disease, the field of neuroscience depends on quantitative studies of neuronal structure and function. Cell morphology and neurite (axonal and dendritic) arborization are typically studied by immunohistochemical and fluorescence techniques. However, dry mass content and intracellular mass transport rates have largely been under-investigated given the inherent difficulties in their measurement. Here, spatial light interference microscopy (SLIM) and dispersion-relation phase spectroscopy (DPS) were used to measure pathlength fluctuations that report on the dry mass and transport within cultured primary neurons across low, medium, and high cell density conditions. It was found that cell density (confluence) affects significantly both the growth rate and mass transport. The analysis method is label-free and does not require neuronal tracing, particle tracking, or neuron reconstruction. Since SLIM can upgrade any existing phase contrast microscope and the imaging and analysis are high-throughput, we anticipate that this approach will be embraced by neuroscientists for broad scale studies. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

  19. Fed-batch strategy for enhancing cell growth and C-phycocyanin production of Arthrospira (Spirulina) platensis under phototrophic cultivation.

    Science.gov (United States)

    Xie, Youping; Jin, Yiwen; Zeng, Xianhai; Chen, Jianfeng; Lu, Yinghua; Jing, Keju

    2015-03-01

    The C-phycocyanin generated in blue-green algae Arthrospira platensis is gaining commercial interest due to its nutrition and healthcare value. In this study, the light intensity and initial biomass concentration were manipulated to improve cell growth and C-phycocyanin production of A.platensis in batch cultivation. The results show that low light intensity and high initial biomass concentration led to increased C-phycocyanin accumulation. The best C-phycocyanin productivity occurred when light intensity and initial biomass concentration were 300μmol/m(2)/s and 0.24g/L, respectively. The fed-batch cultivation proved to be an effective strategy to further enhance C-phycocyanin production of A.platensis. The results indicate that C-phycocyanin accumulation not only requires nitrogen-sufficient condition, but also needs other nutrients. The highest C-phycocyanin content (16.1%), production (1034mg/L) and productivity (94.8mg/L/d) were obtained when using fed-batch strategy with 5mM medium feeding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Comparative Study of Xenobiotic-Free Media for the Cultivation of Human Limbal Epithelial Stem/Progenitor Cells.

    Science.gov (United States)

    González, Sheyla; Chen, Luxia; Deng, Sophie X

    2017-04-01

    The culture of human limbal epithelial stem/progenitor cells (LSCs) in the presence of animal components poses the risk of cross-species contamination in clinical applications. We quantitatively compared different xenobiotic-free culture media for the cultivation of human LSCs. LSCs were cultured from 2 × 2 mm limbal tissue explants on denuded human amniotic membrane with different xenobiotic-free culture media: CnT-Prime (CnT-PR) supplemented with 0%, 1%, 5%, and 10% human serum (HS), embryonic stem cell medium (ESCM) alone or in combination with the standard supplemented hormonal epithelium medium (SHEM, control) at a 1:1 dilution ratio, and modified SHEM (mSHEM), in which cholera toxin and dimethyl sulfoxide (DMSO) were removed, isoproterenol was added, and the epidermal growth factor concentration was reduced. Several parameters were quantified to assess the LSC phenotype: cell morphology, cell growth, cell size, outgrowth size, and expression of the undifferentiated LSC markers cytokeratin (K) 14, and p63α high-expressing (p63α(bright)) cells, a mature keratinocyte marker K12, epithelial marker pancytokeratin (PanK), and stromal cell marker vimentin (Vim). Compared with the standard SHEM control, CnT-PR base medium was associated with a lower cell growth and reduction in the proportion of stem cells generated regardless of the amount of HS supplemented (p  0.05), increased the number of small cells (diameter ≤12 μm; p  0.05). Among all the conditions tested, mSHEM was the most efficient and consistent in supporting the LSC phenotype and growth.

  1. Human Endothelial Cells: Use of Heparin in Cloning and Long-Term Serial Cultivation

    Science.gov (United States)

    Thornton, Susan C.; Mueller, Stephen N.; Levine, Elliot M.

    1983-11-01

    Endothelial cells from human blood vessels were cultured in vitro, with doubling times of 17 to 21 hours for 42 to 79 population doublings. Cloned human endothelial cell strains were established for the first time and had similar proliferative capacities. This vigorous cell growth was achieved by addition of heparin to culture medium containing reduced concentrations of endothelial cell growth factor. The routine cloning and long-term culture of human endothelial cells will facilitate studying the human endothelium in vitro.

  2. Influence of Oxygen in the Cultivation of Human Mesenchymal Stem Cells in Simulated Microgravity: An Explorative Study

    Science.gov (United States)

    Versari, Silvia; Klein-Nulend, Jenneke; van Loon, Jack; Bradamante, Silvia

    2013-02-01

    Previous studies indicated that human Adipose Tissue-derived Mesenchymal Stem Cells (AT-MSCs) cultured in simulated microgravity (sim-μg) in standard laboratory incubators alter their proliferation and differentiation. Recent studies on the stem cell (SC) niches and the influence of oxygen on SC proliferation, senescence, and differentiation point to oxygen level as one of the key regulators of SC fate. Here we present the results of a study that focussed at the evaluation of the influence of oxygen level in the cultivation of AT-MSCs in sim-μg. In detail, cells were cultured for 14 days in sim-μg using the Random Positioning Machine (RPM) and two different oxygen concentrations: 5 % and 20 %. The results were compared with those obtained at 1g in the same conditions. Affymetrix Human Gene 1.0 ST array and Gene Ontology (GO) analysis were performed. The results confirmed that in all of the sim-μg experiments oxygen concentration modulates cell signalling and adhesion, in line with the knowledge that sim-μg affects cell shape and cytoskeletal organization.

  3. Cell density-dependent differential proliferation of neural stem cells on omnidirectional nanopore-arrayed surface.

    Science.gov (United States)

    Cha, Kyoung Je; Kong, Sun-Young; Lee, Ji Soo; Kim, Hyung Woo; Shin, Jae-Yeon; La, Moonwoo; Han, Byung Woo; Kim, Dong Sung; Kim, Hyun-Jung

    2017-10-12

    Recently, the importance of surface nanotopography in the determination of stem cell fate and behavior has been revealed. In the current study, we generated polystyrene cell-culture dishes with an omnidirectional nanopore arrayed surface (ONAS) (diameter: 200 nm, depth: 500 nm, center-to-center distance: 500 nm) and investigated the effects of nanotopography on rat neural stem cells (NSCs). NSCs cultured on ONAS proliferated better than those on the flat surface when cell density was low and showed less spontaneous differentiation during proliferation in the presence of mitogens. Interestingly, NSCs cultured on ONAS at clonal density demonstrated a propensity to generate neurospheres, whereas those on the flat surface migrated out, proliferated as individuals, and spread out to attach to the surface. However, the differential patterns of proliferation were cell density-dependent since the distinct phenomena were lost when cell density was increased. ONAS modulated cytoskeletal reorganization and inhibited formation of focal adhesion, which is generally observed in NSCs grown on flat surfaces. ONAS appeared to reinforce NSC-NSC interaction, restricted individual cell migration and prohibited NSC attachment to the nanopore surface. These data demonstrate that ONAS maintains NSCs as undifferentiated while retaining multipotency and is a better topography for culturing low density NSCs.

  4. Effects of Cultivated Wild Ginseng Herbal Acupuncture to the serum cytokine on Hepatic Metastatic Model using Colon26-L5 Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Byung Jun, Cho

    2006-02-01

    Full Text Available Objectives : This experiment was conducted to evaluate inhibitory effects against hepatic metastasis by cultivated wild ginseng Herbal Acupuncture. Methods : Colon26-L5 carcinoma cells were injected through hepatic portal vein to induce hepatic metastatic cancer. Afer treated cultivated wild ginseng Herbal Acupuncture and investigated various kinds of cytokine level using cytokine chip. Results : 1. Mice treated with cultivated wild ginseng Herbal Acupuncture reduced the level of IL-1α, IL-1β and TNF-α compared to the control group. 2. Mice treated with cultivated wild ginseng Herbal Acupuncture was not showed significant change in the level of IL-4, IL-10, IL-12 and INF-γ compared to the control group. 3. Observing the level of various kinds of cytokine, cultivated wild ginseng Herbal Acupuncture was suppressed pro-inflammatory cytokine. These findings indicate cultivated wild ginseng Herbal Acupuncture is possible to use the inflammatory disease and futher studies carry out for the explanation of anticancer mechanism.

  5. Corneal endothelial cell density and morphology in Phramongkutklao Hospital

    Directory of Open Access Journals (Sweden)

    Narumon Sopapornamorn

    2008-03-01

    Full Text Available Narumon Sopapornamorn1, Manapon Lekskul1, Suthee Panichkul21Department of Ophthalmology, Phramongkutklao Hospital, Bangkok, Thailand; 2Department of Obstetrics and Gynecology, Phramongkutklao College of Medicine, Bangkok, ThailandObjective: To describe the corneal endothelial density and morphology in patients of Phramongkutklao Hospital and the relationship between endothelial cell parameters and other factors.Methods: Four hundred and four eyes of 202 volunteers were included. Noncontact specular microscopy was performed after taking a history and testing the visual acuity, intraocular pressure measurement, Schirmer’s test and routine eye examination by slit lamp microscope. The studied parameters included mean endothelial cell density (MCD, coefficient of variation (CV, and percentage of hexagonality.Results: The mean age of volunteers was 45.73 years; the range being 20 to 80 years old. Their MCD (SD, mean percentage of CV (SD and mean (SD percentage of hexagonality were 2623.49(325 cell/mm2, 39.43(8.23% and 51.50(10.99%, respectively. Statistically, MCD decreased significantly with age (p < 0.01. There was a significant difference in the percentage of CV between genders. There was no statistical significance between parameters and other factors.Conclusion: The normative data of the corneal endothelium of Thai eyes indicated that, statistically, MCD decreased significantly with age. Previous studies have reported no difference in MCD, percentage of CV, and percentage of hexagonality between gender. Nevertheless, significantly different percentages of CV between genders were presented in this study.Keywords: Corneal endothelial cell, parameters, age, gender, smoking, Thailand

  6. Clinical grade cultivation of human Schwann cell, by the using of human autologous serum instead of fetal bovine serum and without growth factors.

    Science.gov (United States)

    Aghayan, Hamid-Reza; Arjmand, Babak; Norouzi-Javidan, Abbas; Saberi, Hooshang; Soleimani, Masoud; Tavakoli, Seyed Amir-Hossein; Khodadadi, Abbas; Tirgar, Niloufar; Mohammadi-Jahani, Fereshteh

    2012-06-01

    Clinical grade cultivation of human schwann cell by the utilization of human autologous serum instead of fetal bovine serum, and also avoiding any growth factors, can increase safety level of this procedure in cases of clinical cell transplantation. The aim of this study was demonstration of the feasibility of clinical grade schwann cell cultivation. In this experimental study after obtaining consent from close relatives we harvested 10 sural nerves from brain death donors and then cultured in 10 seperated culture media plus autologous serum. We also prepared autologous serum from donor's whole blood. Then cultured cells were evaluated by S100 antibody staining for both morphology and purity. Cell purity range was from 97% to 99% (mean=98.11 ± 0.782%). Mean of the cell count was 14,055.56 ± 2,480.479 per micro liter. There was not significant correlation between cell purity and either the culture period or the age of donors (P>0.05). The spearman correlation coefficient for the cell purity with the period or the age of donors was 0.21 and 0.09, respectively. We demonstrated the feasibility of clinical grade schwann cell cultivation by the using of human autologous serum instead of fetal bovine serum and also without the using of growth factors. We also recommended all cell preparation facilities to adhere to the GMP and other similar quality disciplines especially in the preparation of clinically-used cell products.

  7. High power density proton exchange membrane fuel cells

    Science.gov (United States)

    Murphy, Oliver J.; Hitchens, G. Duncan; Manko, David J.

    1993-01-01

    Proton exchange membrane (PEM) fuel cells use a perfluorosulfonic acid solid polymer film as an electrolyte which simplifies water and electrolyte management. Their thin electrolyte layers give efficient systems of low weight, and their materials of construction show extremely long laboratory lifetimes. Their high reliability and their suitability for use in a microgravity environment makes them particularly attractive as a substitute for batteries in satellites utilizing high-power, high energy-density electrochemical energy storage systems. In this investigation, the Dow experimental PEM (XUS-13204.10) and unsupported high platinum loading electrodes yielded very high power densities, of the order of 2.5 W cm(exp -2). A platinum black loading of 5 mg per cm(exp 2) was found to be optimum. On extending the three-dimensional reaction zone of fuel cell electrodes by impregnating solid polymer electrolyte into the electrode structures, Nafion was found to give better performance than the Dow experimental PEM. The depth of penetration of the solid polymer electrolyte into electrode structures was 50-70 percent of the thickness of the platinum-catalyzed active layer. However, the degree of platinum utilization was only 16.6 percent and the roughness factor of a typical electrode was 274.

  8. Assessment of Microbial Fuel Cell Configurations and Power Densities

    KAUST Repository

    Logan, Bruce E.

    2015-07-30

    Different microbial electrochemical technologies are being developed for a many diverse applications, including wastewater treatment, biofuel production, water desalination, remote power sources, and as biosensors. Current and energy densities will always be limited relative to batteries and chemical fuel cells, but these technologies have other advantages based on the self-sustaining nature of the microorganisms that can donate or accept electrons from an electrode, the range of fuels that can be used, and versatility in the chemicals that can be produced. The high cost of membranes will likely limit applications of microbial electrochemical technologies that might require a membrane. For microbial fuel cells, which do not need a membrane, questions remain on whether larger-scale systems can produce power densities similar to those obtained in laboratory-scale systems. It is shown here that configuration and fuel (pure chemicals in laboratory media versus actual wastewaters) remain the key factors in power production, rather than the scale of the application. Systems must be scaled up through careful consideration of electrode spacing and packing per unit volume of reactor.

  9. On-line cell mass monitoring of Saccharomyces cerevisiae cultivations by multi-wavelength fluorescence

    DEFF Research Database (Denmark)

    Haack, Martin Brian; Eliasson, Anna; Olsson, Lisbeth

    2004-01-01

    in a decomposition of the multivariate fluorescent landscape, whereby underlying spectra of the individual intrinsic fluorophors present in the cell mass were estimated. Furthermore, gravimetrically determined cell mass concentration was used together with the fluorescence spectra for calibration and validation...

  10. A protocol for isolation and enriched monolayer cultivation of neural precursor cells from mouse dentate gyrus

    OpenAIRE

    Harish eBabu; Jan-Hendrik eClaasen; Jan-Hendrik eClaasen; Jan-Hendrik eClaasen; Suresh eKannan; Annette E. Rünker; Theo ePalmer; Gerd eKempermann; Gerd eKempermann

    2011-01-01

    In vitro assays are valuable tools to study the characteristics of adult neural precursor cells under controlled conditions with a defined set of parameters. We here present a detailed protocol based on our previous original publication (Babu et al., Enriched monolayer precursor cell cultures from micro-dissected adult mouse dentate gyrus yield functional granule cell-like neurons, PLoS One 2007, 2:e388) to isolate neural precursor cells from the hippocampus of adult mice and maintain and pro...

  11. Nanotransformation of the haemotrophic Mycoplasma suis during in vitro cultivation attempts using modified cell free Mycoplasma media.

    Science.gov (United States)

    Schreiner, Sabrina A; Hoelzle, Katharina; Hofmann-Lehmann, Regina; Hamburger, Anja; Wittenbrink, Max M; Kramer, Manuela M; Sokoli, Albina; Felder, Kathrin M; Groebel, Katrin; Hoelzle, Ludwig E

    2012-11-09

    Mycoplasma suis belongs to haemotrophic mycoplasmas (HMs) which cause infectious anaemia in a large variety of mammals. To date, no in vitro cultivation system for M. suis or other HMs has been established. We hypothesised that M. suis could grow in classical Mycoplasma media supplemented with nutrients (e.g. glucose, iron-binding proteins) which are naturally available from its host environment, the porcine blood. Blood from experimentally M. suis-infected pigs was used to inoculate either standard SP-4 Mycoplasma medium supplemented with iron-binding proteins (transferrin, haemin, and haemoglobin) or glucose-enriched Hayflick Mycoplasma medium. A quantitative M. suis-specific real-time PCR assay was applied to determine and quantify M. suis loads weekly during 12 week-incubation. The first 2 weeks after inoculation M. suis loads decreased remarkably and then persisted at a stationary level over the observation time of 12 weeks in iron-binding protein- or glucose supplemented media variants. Scanning electron microscopic analysis of liquid M. suis sub-cultures on Hayflick agar showed small, densely-packed microcolonies of irregular M. suis cells of reduced size (0.2-0.6μm) indicating nanotransformation. The partial 16S rDNA sequence of these cultured M. suis nanocells was 99.9% identical to M. suis. M. suis cells derived from liquid cultures interact in vitro with porcine erythrocytes by fibril-like structures. We conclude, that the modified Mycoplasma media used for M. suis cultivation are obviously unfavourable for growth but lead to culture persistence. M. suis adapt to inappropriate culture conditions by alteration into nanoforms. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Measuring density and compressibility of white blood cells and prostate cancer cells by microchannel acoustophoresis

    DEFF Research Database (Denmark)

    Barnkob, Rune; Augustsson, Per; Magnusson, Cecilia

    2011-01-01

    We present a novel method for the determination of density and compressibility of individual particles and cells undergoing microchannel acoustophoresis in an arbitrary 2D acoustic field. Our method is a critical advancement within acoustophoretic separation of biological cells, as the ability to...

  13. MDCK and Vero cells for influenza virus vaccine production: a one-to-one comparison up to lab-scale bioreactor cultivation.

    Science.gov (United States)

    Genzel, Yvonne; Dietzsch, Christian; Rapp, Erdmann; Schwarzer, Jana; Reichl, Udo

    2010-09-01

    Over the last decade, adherent MDCK (Madin Darby canine kidney) and Vero cells have attracted considerable attention for production of cell culture-derived influenza vaccines. While numerous publications deal with the design and the optimization of corresponding upstream processes, one-to-one comparisons of these cell lines under comparable cultivation conditions have largely been neglected. Therefore, a direct comparison of influenza virus production with adherent MDCK and Vero cells in T-flasks, roller bottles, and lab-scale bioreactors was performed in this study. First, virus seeds had to be adapted to Vero cells by multiple passages. Glycan analysis of the hemagglutinin (HA) protein showed that for influenza A/PR/8/34 H1N1, three passages were sufficient to achieve a stable new N-glycan fingerprint, higher yields, and a faster increase to maximum HA titers. Compared to MDCK cells, virus production in serum-free medium with Vero cells was highly sensitive to trypsin concentration. Virus stability at 37 degrees C for different virus strains showed differences depending on medium, virus strain, and cell line. After careful adjustment of corresponding parameters, comparable productivity was obtained with both host cell lines in small-scale cultivation systems. However, using these cultivation conditions in lab-scale bioreactors (stirred tank, wave bioreactor) resulted in lower productivities for Vero cells.

  14. Enhanced methane production in an anaerobic digestion and microbial electrolysis cell coupled system with co-cultivation of Geobacter and Methanosarcina.

    Science.gov (United States)

    Yin, Qi; Zhu, Xiaoyu; Zhan, Guoqiang; Bo, Tao; Yang, Yanfei; Tao, Yong; He, Xiaohong; Li, Daping; Yan, Zhiying

    2016-04-01

    The anaerobic digestion (AD) and microbial electrolysis cell (MEC) coupled system has been proved to be a promising process for biomethane production. In this paper, it was found that by co-cultivating Geobacter with Methanosarcina in an AD-MEC coupled system, methane yield was further increased by 24.1%, achieving to 360.2 mL/g-COD, which was comparable to the theoretical methane yield of an anaerobic digester. With the presence of Geobacter, the maximum chemical oxygen demand (COD) removal rate (216.8 mg COD/(L·hr)) and current density (304.3A/m(3)) were both increased by 1.3 and 1.8 fold compared to the previous study without Geobacter, resulting in overall energy efficiency reaching up to 74.6%. Community analysis demonstrated that Geobacter and Methanosarcina could coexist together in the biofilm, and the electrochemical activities of both were confirmed by cyclic voltammetry. Our study observed that the carbon dioxide content in total gas generated from the AD reactor with Geobacter was only half of that generated from the same reactor without Geobacter, suggesting that Methanosarcina may obtain the electron transferred from Geobacter for the reduction of carbon dioxide to methane. Taken together, Geobacter not only can improve the performance of the MEC system, but also can enhance methane production. Copyright © 2015. Published by Elsevier B.V.

  15. In Vitro Protective Effects of Lycium barbarum Berries Cultivated in Umbria (Italy on Human Hepatocellular Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    M. R. Ceccarini

    2016-01-01

    Full Text Available Lycium barbarum is a famous plant in the traditional Chinese medicine. The plant is known to have health-promoting bioactive components. The properties of Lycium barbarum berries cultivated in Umbria (Italy and their effect on human hepatocellular carcinoma cells (HepG2 have been investigated in this work. The obtained results demonstrated that the Lycium barbarum berries from Umbria region display high antioxidant properties evaluated by total phenolic content and ORAC method, on hydrophilic and lipophilic fractions. Moreover, on HepG2 cell line Lycium barbarum berries extract did not change cell viability analyzed by MTT and Trypan blue exclusion assay and did not induce genotoxic effect analyzed by comet assay. Furthermore, it was demonstrated, for the first time, that the berries extract showed a protective effect on DNA damage, expressed as antigenotoxic activity in vitro. Finally, Lycium barbarum berries extract was able to modulate the expression of genes involved in oxidative stress, proliferation, apoptosis, and cancer. In particular, downexpression of genes involved in tumor migration and invasion (CCL5, in increased risk of metastasis and antiapoptotic signal (DUSP1, and in carcinogenesis (GPx-3 and PTGS1, together with overexpression of tumor suppressor gene (MT3, suggested that Umbrian Lycium barbarum berries could play a protective role against hepatocellular carcinoma.

  16. Cultivation in different growth media affects the expression of the cell ...

    African Journals Online (AJOL)

    Environmental factors may greatly influence the expression of cell surface components of bacterial pathogens. Few studies have described the effect of growth conditions on the cell surface hydrophobicity of bacterial isolates of certain Gramnegative and Gram-positive bacteria. The present study describes the effects of ...

  17. Toxoplasma gondii antigens: recovery analysis of tachyzoites cultivated in Vero cell maintained in serum free medium.

    Science.gov (United States)

    da Costa-Silva, Thaís Alves; da Silva Meira, Cristina; Frazzatti-Gallina, Neuza; Pereira-Chioccola, Vera Lucia

    2012-04-01

    Vero cells have been used successfully in Toxoplasma gondii maintenance. Medium supplementation for culture cells with fetal bovine serum is necessary for cellular growth. However, serum in these cultures presents disadvantages, such as the potential to induce hypersensitivity, variability of serum batches, possible presence of contaminants, and the high cost of good quality serum. Culture media formulated without any animal derived components, designed for serum-free growth of cell lines have been used successfully for different virus replication. The advantages of protozoan parasite growth in cell line cultures using serum-free medium remain poorly studied. Thus, this study was designed to determine whether T. gondii tachyzoites grown in Vero cell cultures in serum-free medium, after many passages, are able to maintain the same antigenic proprieties as those maintained in experimental mice. The standardization of Vero cell culture in serum-free medium for in vitro T. gondii tachyzoite production was performed establishing the optimal initial cell concentration for the confluent monolayer formation, which was 1×10(6) Vero cell culture as initial inoculum. The total confluent monolayer formatted after 96 h and the best amount of harvested tachyzoites was 2.1×10(7) using parasite inoculum of 1.5×10(6) after 7 days post-infection. The infectivity of tachyzoites released from Vero cells maintained in serum-free medium was evaluated using groups of Swiss mice infected with cell-culture tachyzoites. The parasite concentrations were similar to those for mice infected with tachyzoites collected from other infected mice. The data from both in vivo and in vitro experiments showed that in at least 30 culture cell passages, the parasites maintained the same infectivity as maintained in vivo. Another question was to know whether in the several continued passages, immunogenic progressive loss could occur. The nucleotide sequences studied were the same between the different

  18. Features of Microsystems for Cultivation and Characterization of Stem Cells with the Aim of Regenerative Therapy

    Directory of Open Access Journals (Sweden)

    Kihoon Ahn

    2016-01-01

    Full Text Available Stem cells have infinite potential for regenerative therapy thanks to their advantageous ability which is differentiable to requisite cell types for recovery and self-renewal. The microsystem has been proved to be more helpful to stem cell studies compared to the traditional methods, relying on its advantageous feature of mimicking in vivo cellular environments as well as other profitable features such as minimum sample consumption for analysis and multiprocedures. A wide variety of microsystems were developed for stem cell studies; however, regenerative therapy-targeted applications of microtechnology should be more emphasized and gain more attractions since the regenerative therapy is one of ultimate goals of biologists and bioengineers. In this review, we introduce stem cell researches harnessing well-known microtechniques (microwell, micropattern, and microfluidic channel in view point of physical principles and how these systems and principles have been implemented appropriately for characterizing stem cells and finding possible regenerative therapies. Biologists may gain information on the principles of microsystems to apply them to find solutions for their current challenges, and engineers may understand limitations of the conventional microsystems and find new chances for further developing practical microsystems. Through the well combination of engineers and biologists, the regenerative therapy-targeted stem cell researches harnessing microtechnology will find better suitable treatments for human disorders.

  19. [Cytogenetic variation of Ungernia victoris cell lines during cultivation on culture media of different composition].

    Science.gov (United States)

    Bublik, E N; Adonin, V I; Kunakh, V A

    2008-01-01

    Chromosome numbers of U. victoris cell lines obtained from the same bulb and cultured for a long time on different agar-solidified and liquid nutrient media differed significantly. The components of the nutrient media including phytohormones did not influence the ratio of cells with different ploidy levels in various lines while transfer of the calluses to the liquid media resulted in the increase of diploid metaphase frequencies.

  20. Endothelial cells on plasma-treated segmented-polyurethane: adhesion strength, antithrombogenicity and cultivation in tubes.

    Science.gov (United States)

    Kawamoto, Y; Nakao, A; Ito, Y; Wada, N; Kaibara, M

    1997-09-01

    When the surface of segmented-polyurethane (SPU), where endothelial cells are not capable of proliferating, is modified by plasma treatment, the adhesion and proliferation of bovine aortic endothelial cells (BAECs) can be drastically improved. The cells were capable of proliferating on the inner surface of a plasma-treated SPU-coated tube (length: 50 mm; inner diameter: 1.5 mm). When a steady flow shear stress of 9 Pa was applied to the cells proliferated on the modified SPU surface for 90 min, most cells did not detach from the surface. From an in vitro evaluation test of antithrombogenicity, the cell surface can be considered to provide an inert surface against thrombus formation and blood coagulation. From analyses of the plasma-treated SPU surface, it was suggested that the improvements in BAEC proliferation and adhesion after plasma treatment were due to the change in wettability of the surface. Data suggest that the plasma treatment would be useful for developing a small-calibre hybrid vascular graft.

  1. Controle genético das células-tronco humanas cultivadas Genetic control of cultivated human stem cells

    Directory of Open Access Journals (Sweden)

    Spencer L. M. Payão

    2009-05-01

    Full Text Available As células-tronco apresentam uma alta capacidade de autorregeneração, assim como, um potencial de diferenciação em uma variedade de tipos celulares. Estas células podem ser classificadas como embrionárias e adultas. Apesar de apresentar propriedades de células-tronco, as mesenquimais apresentam um certo grau de dificuldade no estabelecimento das culturas, podendo induzir a perda da expressão da enzima responsável pela imortalização ou enzima telomerase. A enzima telomerase é considerada um relógio biológico, um indicador que a senescência celular irá se instalar inevitavelmente. A questão mais atual e intrigante dos pesquisadores é se o suposto potencial de divisão, por um determinado período de tempo, das células-tronco cultivadas poderia levar ao acúmulo de alterações genéticas e epigenéticas, resultando em um processo neoplásico. Daí a importância do papel da citogenética humana no controle e monitoramento das células-tronco cultivadas que serão utilizadas na terapia em seres humanos. Alterações cromossômicas estruturais, tais como deleções, translocações e inversões, representam um mecanismo importante pelo qual as células cancerígenas desenvolvem-se gradualmente, uma vez que estas alterações cromossômicas podem levar a uma expressão anormal de muitos genes, podendo desencadear assim o processo neoplásico.Stem cells have a high capacity of self-regeneration, as well as a potential to differentiate into several cell types. These cells can be classified as embryonic or adult. In spite of having inherent properties of stem cells, mesenchymal cells show a certain degree of difficulty to establish cultures. This might induce a loss of the expression of the telomerase enzyme which is considered to be a biological clock or an indicator of the senescence of the cells. The most current and intriguing question for researchers is whether the presumed division potential of cultivated stem cells, over a

  2. The effects of irradiation and microfiltration on the cells growing and total lipids production in the cultivation of Rhodotorula glutinis.

    Science.gov (United States)

    Yen, Hong-Wei; Yang, Ya-Chun

    2012-03-01

    The results of this study indicate that the irradiation could enhance the cells growing of Rhodotorula glutinis to 54.2 ± 1.6g/L as compared to the control (without irradiation) of 38.3 ± 1.2g/L. However, different wavelength of LEDs' (red, green, blue and white) had no significant impacts on the growth and on the lipid content. The accumulation of potential inhibitive metabolic products probably impedes growth, which restricts more biomass accumulated in the fed-batch operation with irradiation. The combining of the fed-batch operation with irradiation and microfiltration can successfully improve the growth of R. glutinis to the maximum of 72.4 ± 0.6g/L and 51.2 ± 4.9% of lipid content obtained. Conclusively, the integration process of a fed-batch operation, irradiation and microfiltration can effectively enhance cell growth in R. glutinis, without any reimbursement of lipid contents. This finding might be useful when applied to the commercialized cultivation of R. glutinis for biodiesel production. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Highly efficient Agrobacterium-mediated transformation of embryogenic cell suspensions of Musa acuminata cv. Mas (AA) via a liquid co-cultivation system.

    Science.gov (United States)

    Huang, Xia; Huang, Xue-Lin; Xiao, Wang; Zhao, Jie-Tang; Dai, Xue-Mei; Chen, Yun-Feng; Li, Xiao-Ju

    2007-10-01

    A high efficient protocol of Agrobacterium-mediated transformation of Musa acuminata cv. Mas (AA), a major banana variety of the South East Asia region, was developed in this study. Male-flower-derived embryogenic cell suspensions (ECS) were co-cultivated in liquid medium with Agrobacterium strain EHA105 harboring a binary vector pCAMBIA2301 carrying nptII and gusA gene in the T-DNA. Depending upon conditions and duration of co-cultivation in liquid medium, 0-490 transgenic plants per 0.5 ml packed cell volume (PCV) of ECS were obtained. The optimum duration of inoculation was 2 h, and the highest transformation frequency was achieved when infected ECS were co-cultivated in liquid medium first for 12 h at 40 rpm and then for 156 h at 100 rpm on a rotary shaker. Co-cultivation for a shorter duration (72 h) or shaking constantly at 100 rpm at the same duration gave 1.6 and 1.8 folds lower transformation efficiency, respectively. No transgenic plants were obtained in parallel experiments carried on semi-solid media. Histochemical GUS assay and molecular analysis in several tissues of the transgenic plants demonstrated that foreign genes were stably integrated into the banana genome. Compared to semi-solid co-cultivation transformation in other banana species, it is remarkable that liquid co-cultivation was much more efficient for transformation of the Mas cultivar, and was at least 1 month faster for regenerating transgenic plants.

  4. Enhancement of cell growth rate by light irradiation in the cultivation of Rhodotorula glutinis.

    Science.gov (United States)

    Yen, Hong-Wei; Zhang, Zhiyong

    2011-10-01

    A yeast, Rhodotorula glutinis, is regarded as a potential microbial oil producer, due to its high lipid content. The flask results of this study indicated that irradiation could increase the growth of R. glutinis compared to that of a batch without irradiation. Further 5-l fermenter results confirmed that irradiation could greatly enhance the cells' growth rate and total lipid productivity. The maximum lipid productivity obtained in the fed-batch operation with 3 LED (light emitting diode) lamps was 0.39 g/l h as compared to 0.34 g/l h in the batch with 3 LED lamps and 0.19 g/l h in the batch without irradiation. Conclusively, the irradiation could significantly increase the cells' growth rate, which, in turn, could be applied to the commercialized production of biodiesel from single cell oils. Copyright © 2011 Elsevier Ltd. All rights reserved.

  5. High-Density Droplet Microarray of Individually Addressable Electrochemical Cells.

    Science.gov (United States)

    Zhang, Huijie; Oellers, Tobias; Feng, Wenqian; Abdulazim, Tarik; Saw, En Ning; Ludwig, Alfred; Levkin, Pavel A; Plumeré, Nicolas

    2017-06-06

    Microarray technology has shown great potential for various types of high-throughput screening applications. The main read-out methods of most microarray platforms, however, are based on optical techniques, limiting the scope of potential applications of such powerful screening technology. Electrochemical methods possess numerous complementary advantages over optical detection methods, including its label-free nature, capability of quantitative monitoring of various reporter molecules, and the ability to not only detect but also address compositions of individual compartments. However, application of electrochemical methods for the purpose of high-throughput screening remains very limited. In this work, we develop a high-density individually addressable electrochemical droplet microarray (eDMA). The eDMA allows for the detection of redox-active reporter molecules irrespective of their electrochemical reversibility in individual nanoliter-sized droplets. Orthogonal band microelectrodes are arranged to form at their intersections an array of three-electrode systems for precise control of the applied potential, which enables direct read-out of the current related to analyte detection. The band microelectrode array is covered with a layer of permeable porous polymethacrylate functionalized with a highly hydrophobic-hydrophilic pattern, forming spatially separated nanoliter-sized droplets on top of each electrochemical cell. Electrochemical characterization of single droplets demonstrates that the underlying electrode system is accessible to redox-active molecules through the hydrophilic polymeric pattern and that the nonwettable hydrophobic boundaries can spatially separate neighboring cells effectively. The eDMA technology opens the possibility to combine the high-throughput biochemical or living cell screenings using the droplet microarray platform with the sequential electrochemical read-out of individual droplets.

  6. Improved poliovirus d-antigen yields by application of different Vero cell cultivation methods

    NARCIS (Netherlands)

    Thomassen, Y.E.; Rubingh, O.; Wijffels, R.H.; Pol, van der L.A.

    2014-01-01

    Vero cells were grown adherent to microcarriers (Cytodex 1; 3 g L-1) using animal component free media in stirred-tank type bioreactors. Different strategies for media refreshment, daily media replacement (semi-batch), continuous media replacement (perfusion) and recirculation of media, were

  7. Co-cultivation of Green Microalgae and Methanotrophic Bacteria for Single Cell Protein Production from Wastewater

    DEFF Research Database (Denmark)

    Rasouli, Zahra; Valverde Pérez, Borja; D'Este, Martina

    2017-01-01

    microalgae – as a means to recover nutrients from industrial wastewater and upcycle them to feed grade single cell protein. Results demonstrated that both algae and bacteria could remove or assimilate most of the organic carbon present in the wastewater. However, their growth stopped before nutrients...

  8. On-line monitoring of responses to nutrient feed additions by multi-frequency permittivity measurements in fed-batch cultivations of CHO cells.

    Science.gov (United States)

    Ansorge, Sven; Esteban, Geoffrey; Schmid, Georg

    2010-04-01

    Changes in the nutrient availability of mammalian cell cultures are reflected in the beta-dispersion parameter characteristic frequency (f ( C )) and the on-line dual frequency permittivity signal. Multi-frequency permittivity measurements were therefore evaluated in fed-batch cultivations of two different CHO cell lines. Similar responses to nutrient depletions and discontinuous feed additions were monitored in different cultivation phases and experimental setups. Sudden increases in permittivity and f ( C ) occurred when feed additions were conducted. A constant or declining permittivity value in combination with a decrease in f ( C ) indicated nutrient limitations. f ( C ) correlated well with changes in oxygen uptake rate when cell diameter remained constant, indicating that metabolic activity is reflected in the value of f ( C ). When significant cell size changes occurred during the cultivations, the analysis of the beta-dispersion parameters was rendered complex. For the application of our findings in other systems it will be hence required to conduct additional off-line measurements. Based on these results, it is hypothesized that multi-frequency permittivity measurements can give information on the intracellular or physiological state in fed-batch mode. Similar observations were made when using different cell lines and feeding strategies, indicating that the findings are transferable to other cell lines and systems. The results should lead to an improved understanding of routine fed-batch processes. Additional studies are, however, required to explore how these observations can be used for fed-batch process development and optimization.

  9. The effect of 193 nm excimer laser radiation on the human corneal endothelial cell density

    Energy Technology Data Exchange (ETDEWEB)

    Isager, P.; Hjortdal, J.Oe.; Ehlers, N. [Aarhus Univ. Hospital, Dept. of Ophthalmology, Aarhus (Denmark)

    1996-06-01

    The effect of 193 nm excimer laser radiation on human corneal endothelial cell density was examined. Fifty-five eyes from 35 patients underwent photorefractive keratectomy for myopia. Photomicrographs of the endothelium were taken a short time before the operation and on an average of 7 months postoperatively with a specular microscope. The average endothelial cell densities were preoperatively 3375 {+-} 266 cells/mm{sup 2} (means {+-} SD) and postoperatively 3348 {+-} 287 cells/mm{sup 2}, corresponding to a fall of 27 cells/mm{sup 2} (N = 55). This fall in endothelial cell density was not statistically significant. A significant correlation between the change in cell density and age of the patient was found, with older patients losing more cells (N = 35, 2p < 0.05). The magnification of the specular microscope was found to change with corneal thickness. The importance of correcting the endothelial cell densities for corneal thickness is discussed. (au) 14 refs.

  10. Cell culture density affects the proliferation activity of human adipose tissue stem cells.

    Science.gov (United States)

    Kim, Dae Seong; Lee, Myoung Woo; Ko, Young Jong; Chun, Yong Hoon; Kim, Hyung Joon; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

    2016-01-01

    In this study, we investigated the effect of cell density on the proliferation activity of human mesenchymal stem cells (MSCs) derived from adipose tissue (AT-MSCs) over time in culture. Passage #4 (P4) and #12 (P12) AT-MSCs from two donors were plated at a density of 200 (culture condition 1, CC1) or 5000 (culture condition 2, CC2) cells cm(-2) . After 7 days of incubation, P4 and P12 AT-MSCs cultured in CC1 were thin and spindle-shaped, whereas those cultured in CC2 had extensive cell-to-cell contacts and an expanded cell volume. In addition, P4 and P12 AT-MSCs in CC1 divided more than three times, while those in CC2 divided less than once on average. Flow cytometric analysis using 5(6)-carboxyfluorescein diacetate N-succinimidyl ester dye showed that the fluorescence intensity of AT-MSCs was lower in CC1 than in CC2. Furthermore, expression of proliferation-associated genes, such as CDC45L, CDC20A and KIF20A, in P4 AT-MSCs was higher in CC1 than in CC2, and this difference was also observed in P12 AT-MSCs. These data demonstrated that cell culture density affects the proliferation activity of MSCs, suggesting that it is feasible to design a strategy to prepare suitable MSCs using specific culture conditions. Copyright © 2016 John Wiley & Sons, Ltd.

  11. Smooth muscle myosin regulation by serum and cell density in cultured rat lung connective tissue cells.

    Science.gov (United States)

    Babij, P; Zhao, J; White, S; Woodcock-Mitchell, J; Mitchell, J; Absher, M; Baldor, L; Periasamy, M; Low, R B

    1993-08-01

    RNA and protein analyses were used to detect expression of SM1 and SM2 smooth muscle myosin heavy chain (MHC) in cultured adult rat lung connective tissue cells (RL-90). Smooth muscle MHC mRNA expression in confluent cells grown in 10% serum was approximately 50% of the level in adult stomach. Similar results were obtained in cells cultured at low density (25% confluency) in 1% serum. However, in low-density cultures transferred to 10% serum for 24 h, the level of MHC mRNA decreased to approximately 20% of that in adult stomach. Smooth muscle alpha-actin showed a pattern of expression similar to that for smooth muscle MHC. Expression of nonmuscle MHC-A mRNA was higher in all culture conditions compared to stomach. MHC-A mRNA expression was less in low-density cultures in low serum and increased when low-density cultures were transferred to 10% serum for 24 h. MHC-B mRNA expression was less in low- vs. high-density cultures. In contrast to MHC-A, however, MHC-B mRNA expression in low-density cultures was higher in low serum. Immunofluorescence and immunoblotting with SM1-specific antibody demonstrated the presence of the SM1 protein isoform as well as reactivity to a protein band migrating slightly faster than SM2. These results demonstrate that cultured rat lung connective tissue cells express smooth muscle MHC and that expression is modulated by culture conditions.

  12. High cell density production of Deinococcus radiodurans under optimized conditions.

    Science.gov (United States)

    He, Yi

    2009-04-01

    Deinococcus radiodurans is a bacterium being investigated for mechanisms of extreme radiation resistance and for bioremediation of environmental radioactive waste sites. In both fundamental and applied research settings, methods for large-scale production of D. radiodurans are needed. In this study, a systematic investigation was carried out to optimize D. radiodurans production at the 20-L fermentor scale. In defined medium, the phosphate buffer typically used was found to be inhibitory to D. radiodurans growth, and caused cell aggregation. Substitution of HEPES and MOPS buffers for phosphate buffer improved D. radiodurans growth characteristics. Several antifoaming agents were investigated to support large-scale production with submerged aeration, and the defoamer KFO 673 was chosen based on its ability to prevent foaming without affecting D. radiodurans growth. The conventional undefined rich medium tryptone/glucose/yeast extract (TGY) maximally supported D. radiodurans growth to an OD(600) of 10. Using a 'design of experiments' approach, we found glucose, Mg and Mn to be critical in supporting high-density growth of D. radiodurans. The optimal pH and temperature for D. radiodurans growth in large-scale preparations were 7.0 and 37 degrees C, respectively. Growth was carried out in a 20-L fermentor using the newly developed media under the optimal conditions. With addition of 10 g/L glucose, 0.5 g/L MgSO(4) . 7H(2)O, 5 microM MnCl(2) into TGY media, an OD(600) of 40 was achieved.

  13. Human endothelial progenitor cells internalize high-density lipoprotein.

    Directory of Open Access Journals (Sweden)

    Kaemisa Srisen

    Full Text Available Endothelial progenitor cells (EPCs originate either directly from hematopoietic stem cells or from a subpopulation of monocytes. Controversial views about intracellular lipid traffic prompted us to analyze the uptake of human high density lipoprotein (HDL, and HDL-cholesterol in human monocytic EPCs. Fluorescence and electron microscopy were used to investigate distribution and intracellular trafficking of HDL and its associated cholesterol using fluorescent surrogates (bodipy-cholesterol and bodipy-cholesteryl oleate, cytochemical labels and fluorochromes including horseradish peroxidase and Alexa Fluor® 568. Uptake and intracellular transport of HDL were demonstrated after internalization periods from 0.5 to 4 hours. In case of HDL-Alexa Fluor® 568, bodipy-cholesterol and bodipy-cholesteryl oleate, a photooxidation method was carried out. HDL-specific reaction products were present in invaginations of the plasma membrane at each time of treatment within endocytic vesicles, in multivesicular bodies and at longer periods of uptake, also in lysosomes. Some HDL-positive endosomes were arranged in form of "strings of pearl"- like structures. HDL-positive multivesicular bodies exhibited intensive staining of limiting and vesicular membranes. Multivesicular bodies of HDL-Alexa Fluor® 568-treated EPCs showed multilamellar intra-vacuolar membranes. At all periods of treatment, labeled endocytic vesicles and organelles were apparent close to the cell surface and in perinuclear areas around the Golgi apparatus. No HDL-related particles could be demonstrated close to its cisterns. Electron tomographic reconstructions showed an accumulation of HDL-containing endosomes close to the trans-Golgi-network. HDL-derived bodipy-cholesterol was localized in endosomal vesicles, multivesicular bodies, lysosomes and in many of the stacked Golgi cisternae and the trans-Golgi-network Internalized HDL-derived bodipy-cholesteryl oleate was channeled into the lysosomal

  14. Estimation of immune cell densities in immune cell conglomerates: an approach for high-throughput quantification.

    Directory of Open Access Journals (Sweden)

    Niels Halama

    Full Text Available BACKGROUND: Determining the correct number of positive immune cells in immunohistological sections of colorectal cancer and other tumor entities is emerging as an important clinical predictor and therapy selector for an individual patient. This task is usually obstructed by cell conglomerates of various sizes. We here show that at least in colorectal cancer the inclusion of immune cell conglomerates is indispensable for estimating reliable patient cell counts. Integrating virtual microscopy and image processing principally allows the high-throughput evaluation of complete tissue slides. METHODOLOGY/PRINCIPAL FINDINGS: For such large-scale systems we demonstrate a robust quantitative image processing algorithm for the reproducible quantification of cell conglomerates on CD3 positive T cells in colorectal cancer. While isolated cells (28 to 80 microm(2 are counted directly, the number of cells contained in a conglomerate is estimated by dividing the area of the conglomerate in thin tissues sections (< or =6 microm by the median area covered by an isolated T cell which we determined as 58 microm(2. We applied our algorithm to large numbers of CD3 positive T cell conglomerates and compared the results to cell counts obtained manually by two independent observers. While especially for high cell counts, the manual counting showed a deviation of up to 400 cells/mm(2 (41% variation, algorithm-determined T cell numbers generally lay in between the manually observed cell numbers but with perfect reproducibility. CONCLUSION: In summary, we recommend our approach as an objective and robust strategy for quantifying immune cell densities in immunohistological sections which can be directly implemented into automated full slide image processing systems.

  15. Umbilical cord blood cells CD133+/CD133- cultivation in neural proliferation media differentiates towards neural cell lineages.

    Science.gov (United States)

    Slovinska, Lucia; Novotna, Ivana; Kubes, Miroslav; Radonak, Jozef; Jergova, Stanislava; Cigankova, Viera; Rosocha, Jan; Cizkova, Dasa

    2011-10-01

    Umbilical cord blood (UCB) has been identified as a good source of hematopoietic and nonhematopoietic stem cells that can be easily isolated. In the present study we investigated the possibility of whether stem cells in mononuclear UCB grown under defined conditions can produce progeny with neural phenotype. A combination of antigen-driven magnetic cell sorting (MACs) method and defined culture conditions specific for cells of neural lineages were used for isolation, expansion and differentiation of CD133+/- cells from UCB. Both UCB-derived fractions were expanded by exposure to growth factors (EGF, bFGF). Differentiation was induced by replacing them with fetal bovine serum. Using immunocytochemistry, the cell markers for neural (MAP2, GFAP, RIP) and non-neural lineages (S-100, von Willebrand factor) were detected. The analysis revealed occurrence of fully mature neural and non-neural lineages, which showed qualitative and quantitative differences between population of CD133+ and CD133- cells. The expression levels of MAP2 and RIP in CD133+ were significantly higher than in CD133-, more GFAP positive cells were found in the CD133-. At the same time, S-100 was expressed by 32.47 ± 6.24% of CD133- cells and 29.42 ± 1.32% of CD133- cell expressed a von Willebrand factor antigen. Our results indicate that stem cells derived from umbilical cord blood are easy to obtain, proliferate and are able to differentiate towards the cells of neural lineages, which represents a promising way for their utilization in cell-based therapies for CNS injuries and diseases. Copyright © 2011 IMSS. Published by Elsevier Inc. All rights reserved.

  16. Poly(3-hydroxybutyrate) anabolism in Cupriavidus necator cultivated at various carbon-to-nitrogen ratios: insights from single-cell Raman spectroscopy

    Science.gov (United States)

    Tao, Zhanhua; Zhang, Pengfei; Qin, Zhaojun; Li, Yong-Qing; Wang, Guiwen

    2016-09-01

    Cupriavidus necator accumulates large amounts of poly(3-hydroxybutyrate) (PHB), a biodegradable substitute for petroleum-based plastics, under certain nutrient conditions. Conventional solvent-extraction-based methods for PHB quantification only obtain average information from cell populations and, thus, mask the heterogeneity among individual cells. Laser tweezers Raman spectroscopy (LTRS) was used to monitor dynamic changes in the contents of PHB, nucleic acids, and proteins in C. necator at the population and single-cell levels when the microorganism cells were cultivated at various carbon-to-nitrogen ratios. The biosynthetic activities of nucleic acids and proteins were maintained at high levels, and only a small amount of PHB was produced when the bacterial cells were cultured under balanced growth conditions. By contrast, the syntheses of nucleic acids and proteins were blocked, and PHB was accumulated in massive amount inside the microbial cells under nitrogen-limiting growth circumstances. Single-cell analysis revealed a relatively high heterogeneity in PHB level at the early stage of the bacterial growth. Additionally, bacterial cells in populations at certain cultivation stages were composed of two or three subpopulations on the basis of their PHB abundance. Overall, LTRS is a reliable single-cell analysis tool that can provide insights into PHB fermentation.

  17. The age-dependent epigenetic and physiological changes in an Arabidopsis T87 cell suspension culture during long-term cultivation

    Energy Technology Data Exchange (ETDEWEB)

    Kwiatkowska, Aleksandra, E-mail: A.Kwiatkows@gmail.com [Department of Botany, University of Rzeszow, Kolbuszowa (Poland); Zebrowski, Jacek [Department of Plant Physiology, University of Rzeszow, Kolbuszowa (Poland); Oklejewicz, Bernadetta [Department of Genetics, University of Rzeszow, Kolbuszowa (Poland); Czarnik, Justyna [Department of Botany, University of Rzeszow, Kolbuszowa (Poland); Halibart-Puzio, Joanna [Department of Plant Physiology, University of Rzeszow, Kolbuszowa (Poland); Wnuk, Maciej [Department of Genetics, University of Rzeszow, Kolbuszowa (Poland)

    2014-05-02

    Highlights: • A decrease in proliferation rate during long-term cultivation of Arabidopsis cells. • Age-dependent increase in senescence-associated gene expression in Arabidopsis cells. • Age-related increase in DNA methylation, H3K9me2, and H3K27me3 in Arabidopsis cells. • High potential of photosynthetic efficiency of long-term cultured Arabidopsis cells. - Abstract: Plant cell suspension cultures represent good model systems applicable for both basic research and biotechnological purposes. Nevertheless, it is widely known that a prolonged in vitro cultivation of plant cells is associated with genetic and epigenetic instabilities, which may limit the usefulness of plant lines. In this study, the age-dependent epigenetic and physiological changes in an asynchronous Arabidopsis T87 cell culture were examined. A prolonged cultivation period was found to be correlated with a decrease in the proliferation rate and a simultaneous increase in the expression of senescence-associated genes, indicating that the aging process started at the late growth phase of the culture. In addition, increases in the heterochromatin-specific epigenetic markers, i.e., global DNA methylation, H3K9 dimethylation, and H3K27 trimethylation, were observed, suggesting the onset of chromatin condensation, a hallmark of the early stages of plant senescence. Although the number of live cells decreased with an increase in the age of the culture, the remaining viable cells retained a high potential to efficiently perform photosynthesis and did not exhibit any symptoms of photosystem II damage.

  18. RNA-binding proteins to assess gene expression states of co-cultivated cells in response to tumor cells

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    Penalva Luiz OF

    2004-09-01

    Full Text Available Abstract Background Tumors and complex tissues consist of mixtures of communicating cells that differ significantly in their gene expression status. In order to understand how different cell types influence one another's gene expression, it will be necessary to monitor the mRNA profiles of each cell type independently and to dissect the mechanisms that regulate their gene expression outcomes. Results In order to approach these questions, we have used RNA-binding proteins such as ELAV/Hu, poly (A binding protein (PABP and cap-binding protein (eIF-4E as reporters of gene expression. Here we demonstrate that the epitope-tagged RNA binding protein, PABP, expressed separately in tumor cells and endothelial cells can be used to discriminate their respective mRNA targets from mixtures of these cells without significant mRNA reassortment or exchange. Moreover, using this approach we identify a set of endothelial genes that respond to the presence of co-cultured breast tumor cells. Conclusion RNA-binding proteins can be used as reporters to elucidate components of operational mRNA networks and operons involved in regulating cell-type specific gene expression in tissues and tumors.

  19. Information in a Network of Neuronal Cells: Effect of Cell Density and Short-Term Depression

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    Valentina Onesto

    2016-01-01

    Full Text Available Neurons are specialized, electrically excitable cells which use electrical to chemical signals to transmit and elaborate information. Understanding how the cooperation of a great many of neurons in a grid may modify and perhaps improve the information quality, in contrast to few neurons in isolation, is critical for the rational design of cell-materials interfaces for applications in regenerative medicine, tissue engineering, and personalized lab-on-a-chips. In the present paper, we couple an integrate-and-fire model with information theory variables to analyse the extent of information in a network of nerve cells. We provide an estimate of the information in the network in bits as a function of cell density and short-term depression time. In the model, neurons are connected through a Delaunay triangulation of not-intersecting edges; in doing so, the number of connecting synapses per neuron is approximately constant to reproduce the early time of network development in planar neural cell cultures. In simulations where the number of nodes is varied, we observe an optimal value of cell density for which information in the grid is maximized. In simulations in which the posttransmission latency time is varied, we observe that information increases as the latency time decreases and, for specific configurations of the grid, it is largely enhanced in a resonance effect.

  20. Information in a Network of Neuronal Cells: Effect of Cell Density and Short-Term Depression

    KAUST Repository

    Onesto, Valentina

    2016-05-10

    Neurons are specialized, electrically excitable cells which use electrical to chemical signals to transmit and elaborate information. Understanding how the cooperation of a great many of neurons in a grid may modify and perhaps improve the information quality, in contrast to few neurons in isolation, is critical for the rational design of cell-materials interfaces for applications in regenerative medicine, tissue engineering, and personalized lab-on-a-chips. In the present paper, we couple an integrate-and-fire model with information theory variables to analyse the extent of information in a network of nerve cells. We provide an estimate of the information in the network in bits as a function of cell density and short-term depression time. In the model, neurons are connected through a Delaunay triangulation of not-intersecting edges; in doing so, the number of connecting synapses per neuron is approximately constant to reproduce the early time of network development in planar neural cell cultures. In simulations where the number of nodes is varied, we observe an optimal value of cell density for which information in the grid is maximized. In simulations in which the posttransmission latency time is varied, we observe that information increases as the latency time decreases and, for specific configurations of the grid, it is largely enhanced in a resonance effect.

  1. Development and optimization of biofilm based algal cultivation

    Science.gov (United States)

    Gross, Martin Anthony

    This dissertation describes research done on biofilm based algal cultivation systems. The system that was developed in this work is the revolving algal biofilm cultivation system (RAB). A raceway-retrofit, and a trough-based pilot-scale RAB system were developed and investigated. Each of the systems significantly outperformed a control raceway pond in side-by-side tests. Furthermore the RAB system was found to require significantly less water than the raceway pond based cultivation system. Lastly a TEA/LCA analysis was conducted to evaluate the economic and life cycle of the RAB cultivation system in comparison to raceway pond. It was found that the RAB system was able to grow algae at a lower cost and was shown to be profitable at a smaller scale than the raceway pond style of algal cultivation. Additionally the RAB system was projected to have lower GHG emissions, and better energy and water use efficiencies in comparison to a raceway pond system. Furthermore, fundamental research was conducted to identify the optimal material for algae to attach on. A total of 28 materials with a smooth surface were tested for initial cell colonization and it was found that the tetradecane contact angle of the materials had a good correlation with cell attachment. The effects of surface texture were evaluated using mesh materials (nylon, polypropylene, high density polyethylene, polyester, aluminum, and stainless steel) with openings ranging from 0.05--6.40 mm. It was found that both surface texture and material composition influence algal attachment.

  2. ULTRASOUND-ENHANCED CELL PRODUCTION OF LACTIC AND PROPIONIC ACID BACTERIA UNDER SUBMERGED CULTIVATION FOR INDUSTRIAL PURPOSES

    Directory of Open Access Journals (Sweden)

    D. A. Durnikin

    2016-08-01

    Full Text Available Lactic and propionic bacteria are actively used as feed and food biopreservatives. The industrial production of these bacteria is carried out using known standard biotechnological approaches and equipment. However, the modern requirements to the volumes of their production require the development of new technologies providing the more intensive growth of bacterial biomass. One of the possible ways to do it is the use of nonspecific stimulators of chemical or physical origin. The stimulating effect of ultrasound on live systems attracts attention of many researchers. Depending on the sonication parameters and conditions, the impact of ultrasound on cell cultures can either stimulate or suppress their life processes. The possibility of the ultrasound stimulation of the biomass accumulation process has been studied for submerged bacterial cultures of Lactococcus lactis VPKM B-2092, Lactobacillus plantarum VPKM B-4173, and Propionibacterium acidipropionici VPKM B-2092. The inoculum with cell contentration of 1 · 108 mL-1 was sonicated at 880 kHz and energy density varied within 0.1-0.7 W/cm3 using a specially designed cuvette, through which the cell suspension was introduced into a fermenter at a rate of 10 mL/s that provided the total sonication time equal to 100-120 seconds. As a signal source, a standard therapeutical ultrasound apparatus UZT-1.01F equipped with a sweep generator was used. For all three cultures, the ultrasound stimulation resulted in a significant increase in the optical density of culture broth comparing to the control and the corresponding increase of the cell concentration. The optimum sonication energy density for the Lactococcus lactis VPKM B-2092, Lactobacillus plantarum VPKM B-4173, and Propionibacterium acidipropionici VPKM B-2092 was equal to 0.5, 0.3-0.5 и 0.3 W/cm3, respectively. Comparing to the control, the cell count of these strains in the culture broth increased in 28.6, 9, and 16.7 times, respectively. Thus

  3. Influence of carvacrol and 1,8-cineole on cell viability, membrane integrity, and morphology of Aeromonas hydrophila cultivated in a vegetable-based broth.

    Science.gov (United States)

    de Sousa, Jossana Pereira; de Oliveira, Kataryne Árabe Rimá; de Figueiredo, Regina Celia Bressan Queiroz; de Souza, Evandro Leite

    2015-02-01

    This study investigated the effects of carvacrol (CAR) and 1,8-cineole (CIN) alone (at the MIC) or in combination at subinhibitory amounts (both at 1/8 MIC) on the cell viability, membrane permeability, and morphology of Aeromonas hydrophila INCQS 7966 (A. hydrophila) cultivated in a vegetable-based broth. CAR and CIN alone or in combination severely affected the viability of the bacteria and caused dramatic changes in the cell membrane permeability, leading to cell death, as observed by confocal laser microscopy. Scanning and transmission electron microscopy images of bacterial cells exposed to CAR or CIN or the mixture of both compounds revealed severe changes in cell wall structure, rupture of the plasma membrane, shrinking of cells, condensation of cytoplasmic content, leakage of intracellular material, and cell collapse. These findings suggest that CAR and CIN alone or in combination at subinhibitory amounts could be applied to inhibit the growth of A. hydrophila in foods, particularly as sanitizing agents in vegetables.

  4. Assessment of the hydration state of sickle cells by phthalate ester density distribution.

    Science.gov (United States)

    Kurantsin-Mills, J; Jacobs, H M; Lessin, L S

    1987-04-01

    Intracellular hemoglobin S (Hb SS) concentration, a function of cell hydration, has a major influence on the rate of Hb SS polymerization and, therefore, cellular sickling. To determine the density distribution of homozygous sickle hemoglobin cells as a function of cell hydration, cells were incubated in autologous plasma buffer mixtures with final osmolalities ranging from 195 to 490 mosm/kg at ambient Po2. The density distribution of the cells was determined by differential flotation on 20 mixtures of di-n-butyl and dimethyl phthalates with specific gravities of 1.062 to 1.142. Mean cell hemoglobin concentration (MCHC) and mean cell volume (MCV) were determined by standard manual procedures. Cell shape was assessed by scanning electron microscopy (SEM), and the axial ratio (L/W) of the elliptical dense cell fraction measured by an image analyzer interfaced with a computer. The density distribution of normal red blood cells lies within a narrow 1.090 to 1.118 gm/ml density band with the middle or transitional 60% (T60) of the cells occupying a density range of 0.0067 +/- 0.0007 gm/ml (+/- SD). The density distribution of sickle cells shows a broader density band of 1.064 to 1.134 gm/ml, and the T60 was 0.0139 +/- 0.0022 gm/ml. The mean T60 did not change with osmotic variation but the mean T60 of Hb SS cells was significantly greater (P less than 0.005). MCHC and 1/MCV varied directly with the median density of the density distribution. By linear regression analysis and Ponder's osmotic equation, it is evident that sickle cells exhibit restricted volume increases in hypotonic media.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. Performance of conversion efficiency of a crystalline silicon solar cell with base doping density

    Directory of Open Access Journals (Sweden)

    Gokhan Sahin

    Full Text Available In this study, we investigate theoretically the electrical parameters of a crystalline silicon solar cell in steady state. Based on a one-dimensional modeling of the cell, the short circuit current density, the open circuit voltage, the shunt and series resistances and the conversion efficiency are calculated, taking into account the base doping density. Either the I-V characteristic, series resistance, shunt resistance and conversion efficiency are determined and studied versus base doping density. The effects applied of base doping density on these parameters have been studied. The aim of this work is to show how short circuit current density, open circuit voltage and parasitic resistances are related to the base doping density and to exhibit the role played by those parasitic resistances on the conversion efficiency of the crystalline silicon solar. Keywords: Crystalline silicon solar cell, Base doping density, Series resistance, Shunt resistance, Conversion efficiency

  6. Collagen Matrix Density Drives the Metabolic Shift in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Brett A. Morris

    2016-11-01

    Full Text Available Increased breast density attributed to collagen I deposition is associated with a 4–6 fold increased risk of developing breast cancer. Here, we assessed cellular metabolic reprogramming of mammary carcinoma cells in response to increased collagen matrix density using an in vitro 3D model. Our initial observations demonstrated changes in functional metabolism in both normal mammary epithelial cells and mammary carcinoma cells in response to changes in matrix density. Further, mammary carcinoma cells grown in high density collagen matrices displayed decreased oxygen consumption and glucose metabolism via the tricarboxylic acid (TCA cycle compared to cells cultured in low density matrices. Despite decreased glucose entry into the TCA cycle, levels of glucose uptake, cell viability, and ROS were not different between high and low density matrices. Interestingly, under high density conditions the contribution of glutamine as a fuel source to drive the TCA cycle was significantly enhanced. These alterations in functional metabolism mirrored significant changes in the expression of metabolic genes involved in glycolysis, oxidative phosphorylation, and the serine synthesis pathway. This study highlights the broad importance of the collagen microenvironment to cellular expression profiles, and shows that changes in density of the collagen microenvironment can modulate metabolic shifts of cancer cells.

  7. Estimation of current density distribution of PAFC by analysis of cell exhaust gas

    Energy Technology Data Exchange (ETDEWEB)

    Kato, S.; Seya, A. [Fuji Electric Co., Ltd., Ichihara-shi (Japan); Asano, A. [Fuji Electric Corporate, Ltd., Yokosuka-shi (Japan)

    1996-12-31

    To estimate distributions of Current densities, voltages, gas concentrations, etc., in phosphoric acid fuel cell (PAFC) stacks, is very important for getting fuel cells with higher quality. In this work, we leave developed a numerical simulation tool to map out the distribution in a PAFC stack. And especially to Study Current density distribution in the reaction area of the cell, we analyzed gas composition in several positions inside a gas outlet manifold of the PAFC stack. Comparing these measured data with calculated data, the current density distribution in a cell plane calculated by the simulation, was certified.

  8. Evaluation of boronate-containing polymer brushes and gels as substrates for carbohydrate-mediated adhesion and cultivation of animal cells.

    Science.gov (United States)

    Ivanov, Alexander E; Kumar, Ashok; Nilsang, Suthasinee; Aguilar, Maria-Rosa; Mikhalovska, Lyubov I; Savina, Irina N; Nilsson, Lars; Scheblykin, Ivan G; Kuzimenkova, Marina V; Galaev, Igor Yu

    2010-02-01

    Boronate-containing thin polyacrylamide gels (B-Gel), polymer brushes (B-Brush) and chemisorbed organosilane layers (B-COSL) were prepared on the surface of glass slides and studied as substrates for carbohydrate-mediated cell adhesion. B-COSL- and B-Brush-modified glass samples exhibited multiple submicron structures densely and irregularly distributed on the glass surface, as found by scanning electron microscopy and atomic force microscopy. B-Gel was ca. 0.1 mm thick and contained pores with effective size of 1-2 microm in the middle and of 5-20 microm on the edges of the gel sample as found by confocal laser scanning microscopy. Evidence for the presence of phenylboronic acid in the samples was given by time-of-flight secondary ion mass-spectrometry (ToF SIMS), contact angle measurements performed in the presence of fructose, and staining with Alizarin Red S dye capable of formation specific, fluorescent complexes with boronic acids. A comparative study of adhesion and cultivation of animal cells on the above substrates was carried out using murine hybridoma M2139 cell line as a model. M2139 cells adhered to the substrates in the culture medium without glucose or sodium pyruvate at pH 8.0, and then were cultivated in the same medium at pH 7.2 for 4 days. It was found that the substrates of B-Brush type were superior both regarding cell adhesion and viability of the adhered cells, among the substrates studied. MTT assay confirmed proliferation of M2139 cells on B-Brush substrates. Some cell adhesion was also registered in the macropores of B-Gel substrate. The effects of surface microstructure of the boronate-containing polymers on cell adhesion are discussed. Transparent glass substrates grafted with boronate-containing copolymers offer good prospects for cell adhesion studies and development of cell-based assays.

  9. A new approach to CAR T-cell gene engineering and cultivation using piggyBac transposon in the presence of IL-4, IL-7 and IL-21.

    Science.gov (United States)

    Ptáčková, Pavlína; Musil, Jan; Štach, Martin; Lesný, Petr; Němečková, Šárka; Král, Vlastimil; Fábry, Milan; Otáhal, Pavel

    2018-02-20

    Clinical-grade chimeric antigenic receptor (CAR)19 T cells are routinely manufactured by lentiviral/retroviral (LV/RV) transduction of an anti-CD3/CD28 activated T cells, which are then propagated in a culture medium supplemented with interleukin (IL)-2. The use of LV/RVs for T-cell modification represents a manufacturing challenge due to the complexity of the transduction approach and the necessity of thorough quality control. We present here a significantly improved protocol for CAR19 T-cell manufacture that is based on the electroporation of peripheral blood mononuclear cells with plasmid DNA encoding the piggyBac transposon/transposase vectors and their cultivation in the presence of cytokines IL-4, IL-7 and IL-21. We found that activation of the CAR receptor by either its cognate ligand (i.e., CD19 expressed on the surface of B cells) or anti-CAR antibody, followed by cultivation in the presence of cytokines IL-4 and IL-7, enables strong and highly selective expansion of functional CAR19 T cells, resulting in >90% CAR + T cells. Addition of cytokine IL-21 to the mixture of IL-4 and IL-7 supported development of immature CAR19 T cells with central memory and stem cell memory phenotypes and expressing very low amounts of inhibitory receptors PD-1, LAG-3 and TIM-3. Our protocol provides a simple and cost-effective method for engineering high-quality T cells for adoptive therapies. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  10. Cultivation of yeast and plant cells entrapped in the low-viscous liquid-core of an alginate membrane capsule prepared using polyethylene glycol.

    Science.gov (United States)

    Koyama, Keitaro; Seki, Minoru

    2004-01-01

    A liquid-core alginate-membrane capsule was prepared by a novel method using polyethylene glycol as a thickener and the cells of Saccharomyces cerevisiae were encapsulated in its core and cultured. After 24 h of cultivation, the cell concentration in the capsule core-liquid reached 222 microg/mm3 on a dry weight basis, which was 1.4 times as large as that in the core of double-layered alginate beads, i.e., alginate-coated alginate-gel beads. The diameter increase of the capsule prepared by the proposed method using immobilized cell growth was suppressed compared to those using the double-layer method and simple alginate-gel bead entrapment, most likely because of the mobility of the entrapped cells in the capsule. We also confirmed that this encapsulation method is applicable for the cultivation of cultured cells of the plant Fragaria ananassa. Additionally, the time-course of the changes in thickener concentration in the liquid-core of the capsule was measured after encapsulation, and revealed the residual thickener, i.e., polyethylene glycol, was able to leak through the alginate shell membrane. This results in low-viscosity of the core liquid enabling good mass-transfer performance, whereas xanthan gum as a thickener could not leak through.

  11. Microfluidic Platform for the Long-Term On-Chip Cultivation of Mammalian Cells for Lab-On-A-Chip Applications.

    Science.gov (United States)

    Bunge, Frank; Driesche, Sander van den; Vellekoop, Michael J

    2017-07-10

    Lab-on-a-Chip (LoC) applications for the long-term analysis of mammalian cells are still very rare due to the lack of convenient cell cultivation devices. The difficulties are the integration of suitable supply structures, the need of expensive equipment like an incubator and sophisticated pumps as well as the choice of material. The presented device is made out of hard, but non-cytotoxic materials (silicon and glass) and contains two vertical arranged membranes out of hydrogel. The porous membranes are used to separate the culture chamber from two supply channels for gases and nutrients. The cells are fed continuously by diffusion through the membranes without the need of an incubator and low requirements on the supply of medium to the assembly. The diffusion of oxygen is modelled in order to find the optimal dimensions of the chamber. The chip is connected via 3D-printed holders to the macroscopic world. The holders are coated with Parlyene C to ensure that only biocompatible materials are in contact with the culture medium. The experiments with MDCK-cells show the successful seeding inside the chip, culturing and passaging. Consequently, the presented platform is a step towards Lab-on-a-Chip applications that require long-term cultivation of mammalian cells.

  12. Cells determine cell density using a small protein bound to a unique tissue-specific phospholipid

    Directory of Open Access Journals (Sweden)

    Christopher J. Petzold

    2013-10-01

    Full Text Available Cell density is the critical parameter controlling tendon morphogenesis. Knowing its neighbors allows a cell to regulate correctly its proliferation and collagen production. A missing link to understanding this process is a molecular description of the sensing mechanism. Previously, this mechanism was shown in cell culture to rely on a diffusible factor (SNZR [sensor] with an affinity for the cell layer. This led to purifying conditioned medium over 4 columns and analyzing the final column fractions for band intensity on SDS gels versus biological activity – a 16 kD band strongly correlated between assays. N-terminal sequencing – EPLAVVDL – identified a large gene (424 AA, extremely conserved between chicken and human. In this paper we probe whether this is the correct gene. Can the predicted large protein be cleaved to a smaller protein? EPLAVVDL occurs towards the C-terminus and cleavage would create a small 94 AA protein. This protein would run at ∼10 kD, so what modifications or cofactor binding accounts for its running at 16 kD on SDS gels? This protein has no prominent hydrophobic regions, so can it be secreted? To validate its role, the chicken cDNA for this gene was tagged with myc and his and transfected into a human osteosarcoma cell line (U2OS. U2OS cells expressed the gene but not passively: differentiating into structures resembling spongy bone and expressing alkaline phosphatase, an early bone marker. Intracellularly, two bands were observed by Western blotting: the full length protein and a smaller form (26 kD. Outside the cell, a small band (28 kD was detected, although it was 40% larger than expected, as well as multiple larger bands. These larger forms could be converted to the predicted smaller protein (94 AA + tags by changing salt concentrations and ultrafiltering – releasing a cofactor to the filtrate while leaving a protein factor in the retentate. Using specific degradative enzymes and mass spectrometry, the

  13. Density, proportion, and dendritic coverage of retinal ganglion cells of the common marmoset (Callithrix jacchus jacchus

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    F.L. Gomes

    2005-06-01

    Full Text Available We performed a quantitative analysis of M and P cell mosaics of the common-marmoset retina. Ganglion cells were labeled retrogradely from optic nerve deposits of Biocytin. The labeling was visualized using horseradish peroxidase (HRP histochemistry and 3-3'diaminobenzidine as chromogen. M and P cells were morphologically similar to those found in Old- and New-World primates. Measurements were performed on well-stained cells from 4 retinas of different animals. We analyzed separate mosaics for inner and outer M and P cells at increasing distances from the fovea (2.5-9 mm of eccentricity to estimate cell density, proportion, and dendritic coverage. M cell density decreased towards the retinal periphery in all quadrants. M cell density was higher in the nasal quadrant than in other retinal regions at similar eccentricities, reaching about 740 cells/mm² at 2.5 mm of temporal eccentricity, and representing 8-14% of all ganglion cells. P cell density increased from peripheral to more central regions, reaching about 5540 cells/mm² at 2.5 mm of temporal eccentricity. P cells represented a smaller proportion of all ganglion cells in the nasal quadrant than in other quadrants, and their numbers increased towards central retinal regions. The M cell coverage factor ranged from 5 to 12 and the P cell coverage factor ranged from 1 to 3 in the nasal quadrant and from 5 to 12 in the other quadrants. These results show that central and peripheral retinal regions differ in terms of cell class proportions and dendritic coverage, and their properties do not result from simply scaling down cell density. Therefore, differences in functional properties between central and peripheral vision should take these distinct regional retinal characteristics into account.

  14. Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus

    OpenAIRE

    Daniel Reyes-Haro; Francisco Emmanuel Labrada-Moncada; Durairaj Ragu Varman; Janina Krüger; Teresa Morales; Ricardo Miledi; Ataúlfo Martínez-Torres

    2016-01-01

    Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA) is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by a...

  15. Density of founder cells affects spatial pattern formation and cooperation in Bacillus subtilis biofilms

    Science.gov (United States)

    van Gestel, Jordi; Weissing, Franz J; Kuipers, Oscar P; Kovács, Ákos T

    2014-01-01

    In nature, most bacteria live in surface-attached sedentary communities known as biofilms. Biofilms are often studied with respect to bacterial interactions. Many cells inhabiting biofilms are assumed to express ‘cooperative traits', like the secretion of extracellular polysaccharides (EPS). These traits can enhance biofilm-related properties, such as stress resilience or colony expansion, while being costly to the cells that express them. In well-mixed populations cooperation is difficult to achieve, because non-cooperative individuals can reap the benefits of cooperation without having to pay the costs. The physical process of biofilm growth can, however, result in the spatial segregation of cooperative from non-cooperative individuals. This segregation can prevent non-cooperative cells from exploiting cooperative neighbors. Here we examine the interaction between spatial pattern formation and cooperation in Bacillus subtilis biofilms. We show, experimentally and by mathematical modeling, that the density of cells at the onset of biofilm growth affects pattern formation during biofilm growth. At low initial cell densities, co-cultured strains strongly segregate in space, whereas spatial segregation does not occur at high initial cell densities. As a consequence, EPS-producing cells have a competitive advantage over non-cooperative mutants when biofilms are initiated at a low density of founder cells, whereas EPS-deficient cells have an advantage at high cell densities. These results underline the importance of spatial pattern formation for competition among bacterial strains and the evolution of microbial cooperation. PMID:24694715

  16. Effect of bone graft density on in vitro cell behavior with enamel matrix derivative.

    Science.gov (United States)

    Miron, Richard J; Caluseru, Oana M; Guillemette, Vincent; Zhang, Yufeng; Buser, Daniel; Chandad, Fatiha; Sculean, Anton

    2015-09-01

    Bone replacement grafting materials play an important role in regenerative dentistry. Despite a large array of tested bone-grafting materials, little information is available comparing the effects of bone graft density on in vitro cell behavior. Therefore, the aim of the present study is to compare the effects of cells seeded on bone grafts at low and high density in vitro for osteoblast adhesion, proliferation, and differentiation. The response of osteoblasts to the presence of a growth factor (enamel matrix derivative, (EMD)) in combination with low (8 mg per well) or high (100 mg per well) bone grafts (BG; natural bone mineral, Bio-Oss®) density, was studied and compared for osteoblast cell adhesion, proliferation, and differentiation as assessed by real-time PCR. Standard tissue culture plastic was used as a control with and without EMD. The present study demonstrates that in vitro testing of bone-grafting materials is largely influenced by bone graft seeding density. Osteoblast adhesion was up to 50 % lower when cells were seeded on high-density BG when compared to low-density BG and control tissue culture plastic. Furthermore, proliferation was affected in a similar manner whereby cell proliferation on high-density BG (100 mg/well) was significantly increased when compared to that on low-density BG (8 mg/well). In contrast, cell differentiation was significantly increased on high-density BG as assessed by real-time PCR for markers collagen 1 (Col 1), alkaline phosphatase (ALP), and osteocalcin (OC) as well as alizarin red staining. The effects of EMD on osteoblast adhesion, proliferation, and differentiation further demonstrated that the bone graft seeding density largely controls in vitro results. EMD significantly increased cell attachment only on high-density BG, whereas EMD was able to further stimulate cell proliferation and differentiation of osteoblasts on control culture plastic and low-density BG when compared to high-density BG. The results

  17. Influência do agrotêxtil sobre a densidade populacional de Monosporascus cannonballus em solo cultivado com melancia (Citrullus lanatus Influence of the row cover on the population density of Monosporascus cannonballus in soil cultivated with watermelon (Citrullus lanatus

    Directory of Open Access Journals (Sweden)

    Erika Valente de Medeiros

    2008-06-01

    Full Text Available O fungo Monosporascus cannonballus é um importante patógeno radicular que ocasiona a síndrome denominada colapso em cucurbitáceas. No presente trabalho, objetivou-se, avaliar os efeitos da cobertura de planta com agrotêxtil branco de 15 g m-2, sobre a densidade populacional de M. cannonballus, em solo cultivado com melancia [Citrullus lanatus (Thumb. Matsu & Nakai]. O delineamento experimental utilizado foi o de blocos casualizados em esquema de parcelas subdivididas, com três repetições. As parcelas foram constituídas por utilização ou não do agrotêxtil e as subparcelas pelas épocas de coleta de solo (0, 12, 24, 36, 48 dias após o transplantio. As variáveis analisadas foram densidade de ascósporos, temperatura do solo e do ar. Não houve influência da utilização da cobertura de agrotêxtil perante a densidade de ascósporos de M. cannonballus, no solo cultivado com melancia, mesmo que tenha proporcionado redução nas temperaturas médias do ar e do solo, sendo uma alternativa eficaz de manejo, na produção da melancia Mickylee.The fungus M. cannonballus is an important root pathogen that causes collapse in cucurbitaceas. The present work had as objective to evaluate the effect of the row cover with white polypropylene of 15 g m-2 on the population density of M.cannonballus in soil cultivated with watermelon [Citrullus lanatus (Thumb. Matsu & Nakai]. The experimental design was of randomized blocks using parcels subdivided scheme with three replications. The parcels consisted of use or not row cover and subparcelas through the time of soil collected (0, 12, 24, 36, 48 days after the transplanting. The variables evaluates were ascospores density, soil temperature and air temperature. There was not influence of the characteristics gotten for the row cover before the ascospores density of M. cannonballus, in the soils cultivated with watermelon and proportioning reduction of soil temperature means and air temperature means

  18. Bulk density of an alfisol under cultivation systems in a long-term experiment evaluated with gamma ray computed tomography;Densidade de um planossolo sob sistemas de cultivo por meio da tomografia computadorizada de raios gama

    Energy Technology Data Exchange (ETDEWEB)

    Bamberg, Adilson Luis; Silva, Thiago Rech da, E-mail: adillbamberg@hotmail.co [Universidade Federal de Pelotas (UFPel), RS (Brazil). Faculdade de Agronomia Eliseu Maciel], E-mail: thiago_cccp@hotmail.com; Pauletto, Eloy Antonio; Pinto, Luiz Fernando Spinelli; Lima, Ana Claudia Rodrigues de, E-mail: pauletto@ufpel.edu.b [Universidade Federal de Pelotas (UFPel), RS (Brazil). Faculdade de Agronomia Eliseu Maciel. Dept. de Solos], E-mail: lfspin@ufpel.edu.b, E-mail: anacrlima@hotmail.co, E-mail: Gome, E-mail: Algenor da Silv, E-mail: algenor@cpact.embrapa.b [EMBRAPA, Pelotas, RS (Brazil). Centro de Pesquisa Agropecuaria Clima Temperado. Estacao Experimental Terras Baixas; Timm, Luis Carlos, E-mail: lctimm@ufpel.edu.b [Universidade Federal de Pelotas (UFPel), RS (Brazil). Faculdade de Agronomia Eliseu Maciel. Dept. de Engenharia Rural

    2009-09-15

    The sustainability of irrigated rice (Oryza sativa L.) in lowland soils is based on the use of crop rotation and succession, which are essential for the control of red and black rice. The effects on the soil properties deserve studies, particularly on soil compaction. The objective of this study was to identify compacted layers in an albaqualf under different cultivation and tillage systems, by evaluating the soil bulk density (Ds) with Gamma Ray Computed Tomography (TC). The analysis was carried out in a long-term experiment, from 1985 to 2004, at an experimental station of EMBRAPA Clima Temperado, Capao do Leao, RS, Brazil, in a random block design with seven treatments, with four replications (T1 - one year rice with conventional tillage followed by two years fallow; T2 - continuous rice under conventional tillage; T4 - rice and soybean (Glycine Max L.) rotation under conventional tillage; T5 - rice, soybean and corn (Zea maize L.) rotation under conventional tillage; T6 - rice under no-tillage in the summer in succession to rye-grass (Lolium multiflorum L.) in the winter; T7 - rice under no-tillage and soybean under conventional tillage rotation; T8 - control: uncultivated soil). The Gamma Ray Computed Tomography method did not identify compacted soil layers under no tillage rice in succession to rye-grass; two fallow years in the irrigated rice production system did not prevent the formation of a compacted layer at the soil surface; and in the rice, soybean and corn rotation under conventional tillage two compacted layers were identified (0.0 to 1.5 cm and 11 to 14 cm), indicating that they may restrict the agricultural production in this cultivation system on Albaqualf soils. (author)

  19. Heterotrophic microalgae cultivation to synergize biodiesel production with waste remediation: progress and perspectives.

    Science.gov (United States)

    Venkata Mohan, S; Rohit, M V; Chiranjeevi, P; Chandra, Rashmi; Navaneeth, B

    2015-05-01

    Microalgae are inexhaustible feedstock for synthesis of biodiesel rich in polyunsaturated fatty acids (PUFA) and valuable bioactive compounds. Their cultivation is critical in sustaining the global economy in terms of human consumption of food and fuel. When compared to autotrophic cultivation, heterotrophic systems are more suitable for producing high cell densities of microalgae for accumulation of large quantities of lipids (triacylglycerols) which can be converted into biodiesel. Consorted efforts are made in this communication to converge recent literature on heterotrophic cultivation systems with simultaneous wastewater treatment and algal oil production. Challenges faced during large scale production and limiting factors which hinder the microalgae growth are enumerated. A strategic deployment of integrated closed loop biorefinery concept with multi-product recovery is proposed to exploit the full potential of algal systems. Sustainable algae cultivation is essential to produce biofuels leading to green future. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Effects of laser peripheral iridotomy on corneal endothelial cell density and cell morphology in primary angle closure suspect subjects

    Directory of Open Access Journals (Sweden)

    Hossein Jamali

    2016-01-01

    Conclusion: In PACS eyes, we did not find a decline in corneal endothelial cell density or a change in cell morphological characteristics, including cell hexagonality and CV, in the central, nasal, and temporal regions of the cornea in any of our subjects over a one-year follow-up period.

  1. Age-related decrease in rod bipolar cell density of the human retina ...

    Indian Academy of Sciences (India)

    During normal ageing, the rods (and other neurones) undergo a significant decrease in density in the human retina from the fourth decade of life onward. Since the rods synapse with the rod bipolar cells in the outer plexiform layer, a decline in rod density (mainly due to death) may ultimately cause an associated decline of ...

  2. Cell density and actomyosin contractility control the organization of migrating collectives within an epithelium

    Science.gov (United States)

    Loza, Andrew J.; Koride, Sarita; Schimizzi, Gregory V.; Li, Bo; Sun, Sean X.; Longmore, Gregory D.

    2016-01-01

    The mechanisms underlying collective migration are important for understanding development, wound healing, and tumor invasion. Here we focus on cell density to determine its role in collective migration. Our findings show that increasing cell density, as might be seen in cancer, transforms groups from broad collectives to small, narrow streams. Conversely, diminishing cell density, as might occur at a wound front, leads to large, broad collectives with a distinct leader–follower structure. Simulations identify force-sensitive contractility as a mediator of how density affects collectives, and guided by this prediction, we find that the baseline state of contractility can enhance or reduce organization. Finally, we test predictions from these data in an in vivo epithelium by using genetic manipulations to drive collective motion between predicted migratory phases. This work demonstrates how commonly altered cellular properties can prime groups of cells to adopt migration patterns that may be harnessed in health or exploited in disease. PMID:27605707

  3. Improving accuracy of cell and chromophore concentration measurements using optical density

    National Research Council Canada - National Science Library

    John A Myers; Brandon S Curtis; Wayne R Curtis

    2013-01-01

      Doc number: 4 Abstract Background: UV-vis spectrophotometric optical density (OD) is the most commonly-used technique for estimating chromophore formation and cell concentration in liquid culture...

  4. Initial cell seeding density influences pancreatic endocrine development during in vitro differentiation of human embryonic stem cells.

    Science.gov (United States)

    Gage, Blair K; Webber, Travis D; Kieffer, Timothy J

    2013-01-01

    Human embryonic stem cells (hESCs) have the ability to form cells derived from all three germ layers, and as such have received significant attention as a possible source for insulin-secreting pancreatic beta-cells for diabetes treatment. While considerable advances have been made in generating hESC-derived insulin-producing cells, to date in vitro-derived glucose-responsive beta-cells have remained an elusive goal. With the objective of increasing the in vitro formation of pancreatic endocrine cells, we examined the effect of varying initial cell seeding density from 1.3 x 10(4) cells/cm(2) to 5.3 x 10(4) cells/cm(2) followed by a 21-day pancreatic endocrine differentiation protocol. Low density-seeded cells were found to be biased toward the G2/M phases of the cell cycle and failed to efficiently differentiate into SOX17-CXCR4 co-positive definitive endoderm cells leaving increased numbers of OCT4 positive cells in day 4 cultures. Moderate density cultures effectively formed definitive endoderm and progressed to express PDX1 in approximately 20% of the culture. High density cultures contained approximately double the numbers of PDX1 positive pancreatic progenitor cells and also showed increased expression of MNX1, PTF1a, NGN3, ARX, and PAX4 compared to cultures seeded at moderate density. The cultures seeded at high density displayed increased formation of polyhormonal pancreatic endocrine cell populations co-expressing insulin, glucagon and somatostatin. The maturation process giving rise to these endocrine cell populations followed the expected cascade of pancreatic progenitor marker (PDX1 and MNX1) expression, followed by pancreatic endocrine specification marker expression (BRN4, PAX4, ARX, NEUROD1, NKX6.1 and NKX2.2) and then pancreatic hormone expression (insulin, glucagon and somatostatin). Taken together these data suggest that initial cell seeding density plays an important role in both germ layer specification and pancreatic progenitor commitment, which

  5. Intratumoral Immune Cell Densities Are Associated with Lung Adenocarcinoma Gene Alterations.

    Science.gov (United States)

    Mansuet-Lupo, Audrey; Alifano, Marco; Pécuchet, Nicolas; Biton, Jérôme; Becht, Etienne; Goc, Jeremy; Germain, Claire; Ouakrim, Hanane; Régnard, Jean-François; Cremer, Isabelle; Laurent-Puig, Pierre; Dieu-Nosjean, Marie-Caroline; Blons, Hélène; Damotte, Diane

    2016-12-01

    Tumor-infiltrating immune cells affect lung cancer outcome. However, the factors that influence the composition and function of the tumor immune environment remain poorly defined and need investigation, particularly in the era of immunotherapy. To determine whether the tumoral immune environment is related to lung adenocarcinoma mutations. This retrospective cohort included 316 consecutive patients with lung adenocarcinoma (225 men; 258 smokers) studied from 2001 to 2005 in a single center. We investigated the association of densities of intratumoral mature dendritic cells (mDCs), CD8(+) T cells, neutrophils, and macrophages with clinical and pathological variables and tumor cell mutation profiles obtained by next-generation sequencing. In 282 tumors, we found 460 mutations, mainly in TP53 (59%), KRAS (40%), STK11 (24%), and EGFR (14%). Intratumoral CD8(+) T-cell density was high in smokers (P = 0.02) and TP53-mutated tumors (P = 0.02) and low in BRAF-mutated tumors (P = 0.005). Intratumoral mDC density was high with low pathological tumor stage (P = 0.01) and low with STK11 mutation (P = 0.004). Intratumoral neutrophil density was high and low with BRAF mutation (P = 0.04) and EGFR mutation (P = 0.02), respectively. Intratumoral macrophage density was low with EGFR mutation (P = 0.01). Intratumoral CD8(+) T-cell and mDC densities remained strong independent markers of overall survival (P = 0.001 and P = 0.02, respectively). Intratumoral immune cell densities (mDCs, CD8(+) T cells, neutrophils, macrophages) were significantly associated with molecular alterations in adenocarcinoma underlying the interactions between cancer cells and their microenvironment.

  6. The relative importance of topography and RGD ligand density for endothelial cell adhesion.

    Directory of Open Access Journals (Sweden)

    Guillaume Le Saux

    Full Text Available The morphology and function of endothelial cells depends on the physical and chemical characteristics of the extracellular environment. Here, we designed silicon surfaces on which topographical features and surface densities of the integrin binding peptide arginine-glycine-aspartic acid (RGD could be independently controlled. We used these surfaces to investigate the relative importance of the surface chemistry of ligand presentation versus surface topography in endothelial cell adhesion. We compared cell adhesion, spreading and migration on surfaces with nano- to micro-scaled pyramids and average densities of 6×10(2-6×10(11 RGD/mm(2. We found that fewer cells adhered onto rough than flat surfaces and that the optimal average RGD density for cell adhesion was 6×10(5 RGD/mm(2 on flat surfaces and substrata with nano-scaled roughness. Only on surfaces with micro-scaled pyramids did the topography hinder cell migration and a lower average RGD density was optimal for adhesion. In contrast, cell spreading was greatest on surfaces with 6×10(8 RGD/mm(2 irrespectively of presence of feature and their size. In summary, our data suggest that the size of pyramids predominately control the number of endothelial cells that adhere to the substratum but the average RGD density governs the degree of cell spreading and length of focal adhesion within adherent cells. The data points towards a two-step model of cell adhesion: the initial contact of cells with a substratum may be guided by the topography while the engagement of cell surface receptors is predominately controlled by the surface chemistry.

  7. Tuning the Density of Poly(ethylene glycol Chains to Control Mammalian Cell and Bacterial Attachment

    Directory of Open Access Journals (Sweden)

    Ahmed Al-Ani

    2017-08-01

    Full Text Available Surface modification of biomaterials with polymer chains has attracted great attention because of their ability to control biointerfacial interactions such as protein adsorption, cell attachment and bacterial biofilm formation. The aim of this study was to control the immobilisation of biomolecules on silicon wafers using poly(ethylene glycol(PEG chains by a “grafting to” technique. In particular, to control the polymer chain graft density in order to capture proteins and preserve their activity in cell culture as well as find the optimal density that would totally prevent bacterial attachment. The PEG graft density was varied by changing the polymer solubility using an increasing salt concentration. The silicon substrates were initially modified with aminopropyl-triethoxysilane (APTES, where the surface density of amine groups was optimised using different concentrations. The results showed under specific conditions, the PEG density was highest with grafting under “cloud point” conditions. The modified surfaces were characterised with X-ray photoelectron spectroscopy (XPS, ellipsometry, atomic force microscopy (AFM and water contact angle measurements. In addition, all modified surfaces were tested with protein solutions and in cell (mesenchymal stem cells and MG63 osteoblast-like cells and bacterial (Pseudomonas aeruginosa attachment assays. Overall, the lowest protein adsorption was observed on the highest polymer graft density, bacterial adhesion was very low on all modified surfaces, and it can be seen that the attachment of mammalian cells gradually increased as the PEG grafting density decreased, reaching the maximum attachment at medium PEG densities. The results demonstrate that, at certain PEG surface coverages, mammalian cell attachment can be tuned with the potential to optimise their behaviour with controlled serum protein adsorption.

  8. Cell death induced by tamoxifen in human blood lymphocytes cultivated in vitro = Morte celular induzida pelo tamoxifeno em linfócitos humanos cultivados in vitro

    Directory of Open Access Journals (Sweden)

    Selma Candelária Genari

    2010-10-01

    Full Text Available Many chemotherapeutic agents with a potential against solid tumors or leukemia can cause lymphopenia. Tamoxifen (TAM is a synthetic non-steroidal anti-estrogen drug employed in female breast cancer treatment. The present study investigated the capacity of TAM to induce cell death in human lymphocytes cultivated in vitro. Lymphocytes were obtained from young (25-30 years; n = 3 and elderly women (58-77 years; n = 3 and cultivated for 24 or 48h, with or without TAM (20 ƒÊM. After the culture, cell viability, immunocytochemical response and ultrastructure were evaluated. TAM affected lymphocytes in a time- dependent manner, and cells obtained from elderly women were the most sensitive to TAM. Immunocytochemicalanalysis evidenced higher frequency of apoptosis in treated cells, and the ultrastructural study revealed autophagic vacuoles, differing from the controls. In summary, the treated lymphocytes were affected by TAM, leading to cell death by apoptosis and autophagy.Muitos agentes quimioterapicos com potencial contra tumores solidos ou leucemias podem causar linfopenia. O Tamoxifeno (TAM e um agente antiestrogeno nao-esteroidal empregado no tratamento de cancer de mama feminino. O presente trabalho investigou a capacidade do TAM em induzir morte celular em linfocitos humanos cultivados in vitro. Oslinfocitos foram obtidos de mulheres jovens (25-30 anos; n = 3 e idosas (58-77 anos; n = 3 e cultivados por 24 ou 48h, com ou sem TAM (20 ƒÊM. Apos a cultura, foram analisadas a viabilidade celular, a resposta imunocitoquimica e a ultraestrutura. Os resultados indicam que o Tamoxifeno induziu morte celular em linfocitos de ambos os grupos, entretanto, as celulas das mulheres idosas apresentaram-se mais sensiveis ao tratamento. A analise imunocitoquimica mostrou maior frequencia de apoptose nas celulas tratadas e o estudo ultraestrutural revelou vacuolos autofagicos nos linfocitos expostos ao Tamoxifeno. Em conclusao, nosso estudo revelou que o TAM

  9. Use of diluted urine for cultivation of Chlorella vulgaris.

    Science.gov (United States)

    Jaatinen, Sanna; Lakaniemi, Aino-Maija; Rintala, Jukka

    2016-01-01

    Our aim was to study the biomass growth of microalga Chlorella vulgaris using diluted human urine as a sole nutrient source. Batch cultivations (21 days) were conducted in five different urine dilutions (1:25-1:300), in 1:100-diluted urine as such and with added trace elements, and as a reference, in artificial growth medium. The highest biomass density was obtained in 1:100-diluted urine with and without additional trace elements (0.73 and 0.60 g L(-1), respectively). Similar biomass growth trends and densities were obtained with 1:25- and 1:300-diluted urine (0.52 vs. 0.48 gVSS L(-1)) indicating that urine at dilution 1:25 can be used to cultivate microalgal based biomass. Interestingly, even 1:300-diluted urine contained sufficiently nutrients and trace elements to support biomass growth. Biomass production was similar despite pH-variation from < 5 to 9 in different incubations indicating robustness of the biomass growth. Ammonium formation did not inhibit overall biomass growth. At the beginning of cultivation, the majority of the biomass consisted of living algal cells, while towards the end, their share decreased and the estimated share of bacteria and cell debris increased.

  10. Multilevel SOT-MRAM Cell with a Novel Sensing Scheme for High-Density Memory Applications

    DEFF Research Database (Denmark)

    Zeinali, Behzad; Madsen, Jens Kargaard; Moradi, Farshad

    in high-density memory application. To deal with this obstacle, we propose a multilevel cell which stores two bits per memory cell. In addition, we propose a novel sensing scheme to read out the stored data in the multilevel SOT-MRAM cell. Our simulation results show that the proposed cell can achieve 3X......This paper presents a multilevel spin-orbit torque magnetic random access memory (SOT-MRAM). The conventional SOT-MRAMs enables a reliable and energy efficient write operation. However, these cells require two access transistors per cell, hence the efficiency of the SOTMRAMs can be questioned...

  11. Insulin-like growth factor-II receptors in cultured rat hepatocytes: regulation by cell density

    Energy Technology Data Exchange (ETDEWEB)

    Scott, C.D.; Baxter, R.C.

    1987-12-01

    Insulin-like growth factor-II (IGF-II) receptors in primary cultures of adult rat hepatocytes were characterized and their regulation by cell density examined. In hepatocytes cultured at 5 X 10(5) cells per 3.8 cm2 plate (/sup 125/I)IGF-II bound to specific, high affinity receptors (Ka = 4.4 +/- 0.5 X 10(9) l/mol). Less than 1% cross-reactivity by IGF-I and no cross-reactivity by insulin were observed. IGF-II binding increased when cells were permeabilized with 0.01% digitonin, suggesting the presence of an intracellular receptor pool. Determined by Scatchard analysis and by polyacrylamide gel electrophoresis after affinity labeling, the higher binding was due solely to an increase in binding sites present on 220 kDa type II IGF receptors. In hepatocytes cultured at low densities, the number of cell surface receptors increased markedly, from 10-20,000 receptors per cell at a culture density of 6 X 10(5) cells/well to 70-80,000 receptors per cell at 0.38 X 10(5) cells/well. The increase was not due simply to the exposure of receptors from the intracellular pool, as a density-related increase in receptors was also seen in cells permeabilized with digitonin. There was no evidence that IGF binding proteins, either secreted by hepatocytes or present in fetal calf serum, had any effect on the measurement of receptor concentration or affinity. We conclude that rat hepatocytes in primary culture contain specific IGF-II receptors and that both cell surface and intracellular receptors are regulated by cell density.

  12. Expansion of adipose mesenchymal stromal cells is affected by human platelet lysate and plating density.

    Science.gov (United States)

    Cholewa, Dominik; Stiehl, Thomas; Schellenberg, Anne; Bokermann, Gudrun; Joussen, Sylvia; Koch, Carmen; Walenda, Thomas; Pallua, Norbert; Marciniak-Czochra, Anna; Suschek, Christoph V; Wagner, Wolfgang

    2011-01-01

    The composition of mesenchymal stromal cells (MSCs) changes in the course of in vitro culture expansion. Little is known how these cell preparations are influenced by culture media, plating density, or passaging. In this study, we have isolated MSCs from human adipose tissue in culture medium supplemented with either fetal calf serum (FCS) or human platelet lysate (HPL). In addition, culture expansion was simultaneously performed at plating densities of 10 or 10,000 cells/cm(2). The use of FCS resulted in larger cells, whereas HPL significantly enhanced proliferation. Notably, HPL also facilitated expansion for more population doublings than FCS (43 ± 3 vs. 22 ± 4 population doubling; p < 0.001), while plating density did not have a significant effect on long-term growth curves. To gain further insight into population dynamics, we conceived a cellular automaton model to simulate expansion of MSCS. It is based on the assumptions that the number of cell divisions is limited and that due to contact inhibition proliferation occurs only at the rim of colonies. The model predicts that low plating densities result in more heterogeneity with regard to cell division history, and favor subpopulations of higher migratory activity. In summary, HPL is a suitable serum supplement for isolation of MSC from adipose tissue and facilitates more population doublings than FCS. Cellular automaton computer simulations provided additional insights into how complex population dynamics during long-term expansion are affected by plating density and migration.

  13. Classification of cultivated plants.

    NARCIS (Netherlands)

    Brandenburg, W.A.

    1986-01-01

    Agricultural practice demands principles for classification, starting from the basal entity in cultivated plants: the cultivar. In establishing biosystematic relationships between wild, weedy and cultivated plants, the species concept needs re-examination. Combining of botanic classification, based

  14. Effects of cultivation conditions and media composition on cell growth and lipid productivity of indigenous microalga Chlorella vulgaris ESP-31.

    Science.gov (United States)

    Yeh, Kuei-Ling; Chang, Jo-Shu

    2012-02-01

    The growth and lipid productivity of an isolated microalga Chlorella vulgaris ESP-31 were investigated under different media and cultivation conditions, including phototrophic growth (NaHCO(3) or CO(2), with light), heterotrophic growth (glucose, without light), photoheterotrophic growth (glucose, with light) and mixotrophic growth (glucose and CO(2), with light). C. vulgaris ESP-31 preferred to grow under phototrophic (CO(2)), photoheterotrophic and mixotrophic conditions on nitrogen-rich medium (i.e., Basal medium and Modified Bristol's medium), reaching a biomass concentration of 2-5 g/l. The growth on nitrogen-limiting MBL medium resulted in higher lipid accumulation (20-53%) but slower growth rate. Higher lipid content (40-53%) and higher lipid productivity (67-144 mg/l/d) were obtained under mixotrophic cultivation with all the culture media used. The fatty acid composition of the microalgal lipid comprises over 60-68% of saturated fatty acids (i.e., palmitic acid (C16:0), stearic acid (C18:0)) and monounsaturated acids (i.e., oleic acid (C18:1)). This lipid composition is suitable for biodiesel production. Copyright © 2011 Elsevier Ltd. All rights reserved.

  15. Effect of Cell Seeding Density and Inflammatory Cytokines on Adipose Tissue-Derived Stem Cells: an in Vitro Study.

    Science.gov (United States)

    Sukho, Panithi; Kirpensteijn, Jolle; Hesselink, Jan Willem; van Osch, Gerjo J V M; Verseijden, Femke; Bastiaansen-Jenniskens, Yvonne M

    2017-04-01

    Adipose tissue-derived stem cells (ASCs) are known to be able to promote repair of injured tissue via paracrine factors. However, the effect of cell density and inflammatory cytokines on the paracrine ability of ASCs remains largely unknown. To investigate these effects, ASCs were cultured in 8000 cells/cm2, 20,000 cells/cm2, 50,000 cells/cm2, and 400,000 cells/cm2 with and without 10 or 20 ng/ml tumor necrosis factor alpha (TNFα) and 25 or 50 ng/ml interferon gamma (IFNγ). ASC-sheets formed at 400,000 cells/cm2 after 48 h of culture. With increasing concentrations of TNFα and IFNγ, ASC-sheets with 400,000 cells/cm2 had increased production of angiogenic factors Vascular Endothelial Growth Factor and Fibroblast Growth Factor and decreased expression of pro-inflammatory genes TNFA and Prostaglandin Synthase 2 (PTGS2) compared to lower density ASCs. Moreover, the conditioned medium of ASC-sheets with 400,000 cells/cm2 stimulated with the low concentration of TNFα and IFNγ enhanced endothelial cell proliferation and fibroblast migration. These results suggest that a high cell density enhances ASC paracrine function might beneficial for wound repair, especially in pro-inflammatory conditions.

  16. Investigation of expression and activity levels of primary rat hepatocyte detoxication genes under various flow rates and cell densities in microfluidic biochips.

    Science.gov (United States)

    Baudoin, Régis; Alberto, Giulia; Legendre, Audrey; Paullier, Patrick; Naudot, Marie; Fleury, Marie-José; Jacques, Sébastien; Griscom, Laurent; Leclerc, Eric

    2014-01-01

    We investigated the behavior of primary rat hepatocytes in biochips using a microfluidic platform (the integrated dynamic cell culture microchip). We studied the effects of cell inoculation densities (0.2-0.5 × 10(6) cells/biochip) and perfusion flow rates (10, 25, and 40 µL/min) during 72 h of perfusion. No effects were observed on hepatocyte morphology, but the levels of mRNA and CYP1A2 activity were found to be dependent on the initial cell densities and flow rates. The dataset made it possible to extract a best estimated range of parameters in which the rat hepatocytes appeared the most functional in the biochips. Namely, at 0.25 × 10(6) inoculated cells cultivated at 25 µL/min for 72 h, we demonstrated better induction of the expression of all the genes analyzed in comparison with other cell densities and flow rates. More precisely, when primary rat hepatocytes were cultivated at these conditions, the time-lapse analysis demonstrated an over expression of CYP3A1, CYP2B1, ABCC1b and ABCC2 in the biochips when compared to the postextraction levels. Furthermore, the AHR, CYP1A2, GSTA2, SULT1A1, and UGT1A6 levels remained higher than 50% of the postextraction values whereas values of HNF4α, CEBP, and PXR remained higher than 20% during the duration of the culture process. Nevertheless, an important reduction in mRNA levels was found for the xenosensors CAR and FXR, and the related CYP (CYP2E1, CYP7A1, CYP3A2, and CYP2D2). CYP1A2 functionality was illustrated by 700 ± 100 pmol/h/10(6) cells resorufin production. This study highlighted the functionality in optimized conditions of primary rat hepatocytes in parallelized microfluidic cultures and their potential for drug screening applications. © 2013 American Institute of Chemical Engineers.

  17. CELL-DENSITY MODULATES GROWTH, EXTRACELLULAR-MATRIX, AND PROTEIN-SYNTHESIS OF CULTURED RAT MESANGIAL CELLS

    NARCIS (Netherlands)

    WOLTHUIS, A; BOES, A

    1993-01-01

    Mesangial cell (MC) hyperplasia and accumulation of extracellular matrix are hallmarks of chronic glomerular disease. The present in vitro study examined the effects of cell density on growth, extracellular matrix formation, and protein synthesis of cultured rat MCs. A negative linear relationship

  18. Microfluidic Adaptation of Density-Gradient Centrifugation for Isolation of Particles and Cells

    Directory of Open Access Journals (Sweden)

    Yuxi Sun

    2017-08-01

    Full Text Available Density-gradient centrifugation is a label-free approach that has been extensively used for cell separations. Though elegant, this process is time-consuming (>30 min, subjects cells to high levels of stress (>350 g and relies on user skill to enable fractionation of cells that layer as a narrow band between the density-gradient medium and platelet-rich plasma. We hypothesized that microfluidic adaptation of this technique could transform this process into a rapid fractionation approach where samples are separated in a continuous fashion while being exposed to lower levels of stress (<100 g for shorter durations of time (<3 min. To demonstrate proof-of-concept, we designed a microfluidic density-gradient centrifugation device and constructed a setup to introduce samples and medium like Ficoll in a continuous, pump-less fashion where cells and particles can be exposed to centrifugal force and separated via different outlets. Proof-of-concept studies using binary mixtures of low-density polystyrene beads (1.02 g/cm3 and high-density silicon dioxide beads (2.2 g/cm3 with Ficoll–Paque (1.06 g/cm3 show that separation is indeed feasible with >99% separation efficiency suggesting that this approach can be further adapted for separation of cells.

  19. Induction of xylanases by sugar cane bagasse at different cell densities of Cellulomonas flavigena.

    Science.gov (United States)

    Amaya-Delgado, L; Vega-Estrada, J; Flores-Cotera, L B; Dendooven, L; Hidalgo-Lara, M E; Montes-Horcasitas, M C

    2006-04-01

    The effect of cell density on xylanolytic activity and productivity of Cellulomonas flavigena was evaluated under two different culturing conditions: fed-batch culture with discontinuous feed of sugar cane bagasse (SCB; condition 1) and glycerol fed-batch culture followed by addition of SBC as xylanases inducer (condition 2). The enzymatic profile of xylanases was similar in both systems, regardless of the initial cell density at time of induction. However, the xylanolytic activity changed with initial cell density at the time of induction (condition 2). The maximum volumetric xylanase activity increased with increased initial cell density from 4 to 34 g l(-1) but decreased above this value. The largest total volumetric xylanase productivity under condition 2 (1.3 IU ml(-1) h(-1)) was significantly greater compared to condition 1 (maximum 0.6 IU ml(-1) h(-1)). Consequently, induction of xylanase activity by SCB after growing of C. flavigena on glycerol at intermediate cell density can be a feasible alternative to improve activity and productivity of xylanolytic enzymes.

  20. Resistance of human brain microvascular endothelial cells in culture to methylmercury: cell-density-dependent defense mechanisms.

    Science.gov (United States)

    Hirooka, Takashi; Fujiwara, Yasuyuki; Shinkai, Yasuhiro; Yamamoto, Chika; Yasutake, Akira; Satoh, Masahiko; Eto, Komyo; Kaji, Toshiyuki

    2010-06-01

    Vascular toxicity is important for understanding the neurotoxicity of methylmercury, because microvessels strongly influence the construction of microenvironment around neurons. Previously, we found that low density-human brain microvascular pericytes are markedly susceptible to methylmercury cytotoxicity due to high expression levels of the L-type amino acid transporter 1 (LAT-1) that transports methylmercury into the cells. Although LAT-1 can be, in general, highly expressed in sparse cells that require amino acids for growth, we found that human brain microvascular endothelial cells, regardless of cell density, were resistant to methylmercury cytotoxicity. To investigate the mechanisms underlying this resistance, we exposed the endothelial cells at low and high cell densities to methylmercury and determined the extent of nonspecific cell damage, intracellular accumulation of methylmercury, expression of LAT-1 and LAT-2 mRNAs, and intracellular expression of reduced glutathione and metallothionein. These experiments indicate that sparse endothelial cells intracellularly accumulate more methylmercury via the highly expressed LAT-1, but are resistant to methylmercury cytotoxicity by higher expression of the protective sulfhydryl peptides, namely, reduced glutathione and metallothionein. It is suggested that both nonspecific and functional damage is caused in pericytes, whereas functional abnormalities rather than nonspecific damage may occur to a greater extent in the endothelial cells in the brain microvessels exposed to methylmercury. The previous and present data also suggest that methylmercury exhibits toxicity in endothelial cells in a manner different from that in pericytes in the brain microvessels.

  1. Modelling and simulation of double chamber microbial fuel cell. Cell voltage, power density and temperature variation with process parameters

    Energy Technology Data Exchange (ETDEWEB)

    Shankar, Ravi; Mondal, Prasenjit; Chand, Shri [Indian Institute of Technology Roorkee, Uttaranchal (India). Dept. of Chemical Engineering

    2013-11-01

    In the present paper steady state models of a double chamber glucose glutamic acid microbial fuel cell (GGA-MFC) under continuous operation have been developed and solved using Matlab 2007 software. The experimental data reported in a recent literature has been used for the validation of the models. The present models give prediction on the cell voltage and cell power density with 19-44% errors, which is less (up to 20%) than the errors on the prediction of cell voltage made in some recent literature for the same MFC where the effects of the difference in pH and ionic conductivity between anodic and cathodic solutions on cell voltage were not incorporated in model equations. It also describes the changes in anodic and cathodic chamber temperature due to the increase in substrate concentration and cell current density. Temperature profile across the membrane thickness has also been studied. (orig.)

  2. Growth and lipid synthesis promotion in mixotrophic Neochloris oleoabundans (Chlorophyta) cultivated with glucose.

    Science.gov (United States)

    Giovanardi, Martina; Baldisserotto, Costanza; Ferroni, Lorenzo; Longoni, Paolo; Cella, Rino; Pancaldi, Simonetta

    2014-01-01

    In the recent years, the studies concerning the cultivation of Neochloris oleoabundans for biofuel purposes have increased, in relation to its capability to accumulate lipids when grown under nutrient starvation. Unfortunately, this cultivation mode does not allow to reach high biomass densities, which are required to improve the feasibility of the process. Increasing knowledge of the microalgal physiology is necessary to obtain new useful information for the improvement of culture performance in the perspective of large-scale cultivation. In this work, the mixotrophic cultivation of N. oleoabundans in a brackish medium added with different glucose concentrations has been tested under shaking, with the aim of stimulating growth alongside lipid accumulation inside cells. Cell morphology, glucose consumption, photosynthetic pigment content and photosynthetic efficiency were also investigated. Among all tested glucose concentrations (0-30 g L(-1)), it was observed that 2.5 g L(-1) was the optimal concentration, allowing to obtain the best compromise between glucose supplement, biomass production and lipid accumulation. Growth was highly enhanced in mixotrophic cultures, linked to the release of cells from sporocysts. A unique feature characterising mixotrophy in N. oleoabundans was the promotion of the maximum quantum yield of Photosystem II. Moreover, when mixotrophic cells entered the stationary phase, high lipid accumulation was induced. This study shows that the addition of glucose to N. oleoabundans remarkably increases the production of biomass enriched in lipids and represents an advancement for the cultivation of this microalga for applied purposes.

  3. Immunomodulating activities of cultivated maitake medicinal mushroom Grifola frondosa (Dicks.: Fr.) S.F. Gray (higher Basidiomycetes) on peripheral blood mononuclear cells.

    Science.gov (United States)

    Svagelj, Mirjan; Berovic, Marin; Gregori, Andrej; Wraber, Branka; Simcic, Sasa; Boh, Bojana

    2012-01-01

    Grifola frondosa is a culinary-medicinal mushroom that contains several physiologically active compounds, of which polysaccharides, specifically β-glucans, are known to possess immunomodulating properties. Its extracts are studied for application as adjuncts for chemotherapy, and experiments in animal models support the use of this mushroom for cancer treatment. The effect of extracts obtained from mushrooms cultivated on different substrates and their capacity of inducing the secretion of cytokines from human peripheral blood mononuclear cells were studied. The activity of extracts at concentrations 12.5, 100, and 200 μg/mL on induction of TNF-α, IFN-γ, and IL-12 was screened. Two extracts from substrates fortified with olive oil press cakes showed appreciable activity and induced the secretion of TNF-α, IL-12, and INF-γ. The extracts differed from the others in the amount of sugar, protein, and β-glucans, which can explain their higher activity. Present results show that different substrates and different source materials can reasonably modify the bioactivity of cultivated G. frondosa.

  4. Intrinsic cardiac adrenergic (ICA) cell density and MAO-A activity in failing rat hearts.

    Science.gov (United States)

    van Eif, Vincent W W; Bogaards, Sylvia J P; van der Laarse, Willem J

    2014-02-01

    The efficiency (work/oxygen consumption) of isolated papillary muscles from failing hearts is reduced. We investigated whether this can be due to an increase of intrinsic cardiac adrenergic (ICA) cell density. The number of ICA cells in the septum and both ventricular walls was determined by tyrosine hydroxylase immunohistochemistry in rats with monocrotaline-induced pulmonary hypertension. We found that the number of ICA cells is about 200,000 per rat heart. ICA cell density was significantly lower in right ventricular myocardium of hypertrophied hearts (P ICA cell density and MAO-A activity was absent. Clorgyline (2 μM) decreased the basal rate of oxygen consumption of right ventricular papillary muscles by 65 μM O(2)/s (P = 0.027). This rate can only be maintained for several seconds judging from the catecholamine content of the preparations reported previously. High ICA cell activity rather than density and/or recycling of oxidized catecholamines are discussed as alternative explanations for the low myocardial efficiency in experimental pulmonary hypertension.

  5. Epithelial cell density in cataractous lenses of patients with diabetes: association with erythrocyte aldose reductase.

    Science.gov (United States)

    Kumamoto, Yuko; Takamura, Yoshihiro; Kubo, Eri; Tsuzuki, Syousai; Akagi, Yoshio

    2007-09-01

    In the present study we evaluated the cell density of lens epithelium and its relation to the degree of erythrocyte aldose reductase (AR) in patients with type 2 diabetes. This prospective clinical study included 46 eyes of patients with type 2 diabetes and 48 eyes of patients without diabetes mellitus (DM). Flat preparations of lens epithelial cells (LECs) attached to the anterior capsule were studied. Multiple regression analysis was performed to evaluate the association between lens cell density and age, gender, type of cataract, duration of diabetes, diabetic retinopathy (DR), the levels of glycosylated hemoglobin (HbA1c) and erythrocyte AR. The mean density of LECs of patients with type 2 diabetes was 4,141+/-508cells/mm(2), which was significantly lower than that of patients without DM (4,560+/-458cells/mm(2); p6.5%) or with DR. These results suggest that the polyol pathway via AR may be associated with the reduction of epithelial cell density in the eyes of patients with DM.

  6. Tissue non-specific alkaline phosphatase production by human dental pulp stromal cells is enhanced by high density cell culture.

    Science.gov (United States)

    Tomlinson, Matthew J; Dennis, Caitriona; Yang, Xuebin B; Kirkham, Jennifer

    2015-08-01

    The cell surface hydrolase tissue non-specific alkaline phosphatase (TNAP) (also known as MSCA-1) is used to identify a sub-population of bone marrow stromal cells (BMSCs) with high mineralising potential and is found on subsets of cells within the dental pulp. We aim to determine whether TNAP is co-expressed by human dental pulp stromal cells (hDPSCs) alongside a range of BMSC markers, whether this is an active form of the enzyme and the effects of culture duration and cell density on its expression. Cells from primary dental pulp and culture expanded hDPSCs expressed TNAP. Subsequent analyses revealed persistent TNAP expression and co-expression with BMSC markers such as CD73 and CD90. Flow cytometry and biochemical assays showed that increased culture durations and cell densities enhanced TNAP expression by hDPSCs. Arresting the hDPSC cell cycle also increased TNAP expression. These data confirm that TNAP is co-expressed by hDPSCs together with other BMSC markers and show that cell density affects TNAP expression levels. We conclude that TNAP is a potentially useful marker for hDPSC selection especially for uses in mineralised tissue regenerative therapies.

  7. Microstructure characterisation of solid oxide electrolysis cells operated at high current density

    DEFF Research Database (Denmark)

    Bowen, Jacob R.; Bentzen, Janet Jonna; Chen, Ming

    High temperature solid oxide cells can be operated either as fuel cells or electrolysis cells for efficient power generation or production of hydrogen from steam or synthesis gas (H2 + CO) from steam and CO2 respectively. When operated under harsh conditions, they often exhibit microstructural......, microstructure evolution of the Ni-yttria stabilized zirconia (YSZ) is followed as a function of galvanostatic steam electrolysis testing at current densities between -0.5 and -1.0 A cm-2 for periods of up to 750 hours at 800 °C. The volume fraction and size of the percolating Ni particles was statistically...... quantified using the mean linear intercept method as a function of current density and correlated to increases in serial resistance. The above structural changes are then compared in terms of electrode degradation observed during the co-electrolysis of steam and CO2 at current densities up to -1.5 A cm-2...

  8. Cell damage from radiation-induced bystander effects for different cell densities simulated by a mathematical model via cellular automata

    Energy Technology Data Exchange (ETDEWEB)

    Meireles, Sincler P. de; Santos, Adriano M.; Grynberg, Suely Epsztein, E-mail: spm@cdtn.b, E-mail: amsantos@cdtn.b, E-mail: seg@cdtn.b [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil); Nunes, Maria Eugenia S., E-mail: mariaeugenia@iceb.ufop.b [Universidade Federal de Ouro Preto (UFOP), MG (Brazil)

    2011-07-01

    During recent years, there has been a shift from an approach focused entirely on DNA as the main target of ionizing radiation to a vision that considers complex signaling pathways in cells and among cells within tissues. Several newly recognized responses were classified as the so-called non-target responses in which the biological effects are not directly related to the amount of energy deposited in the DNA of cells that were traversed by radiation. In 1992 the bystander effect was described referring to a series of responses such as death, chromosomal instability or other abnormalities that occur in non-irradiated cells that came into contact with irradiated cells or medium from irradiated cells. In this work, we have developed a mathematical model via cellular automata, to quantify cell death induced by the bystander effect. The model is based on experiments with irradiated cells conditioned medium which suggests that irradiated cells secrete molecules in the medium that are capable of damaging other cells. The computational model consists of two-dimensional cellular automata which is able to simulate the transmission of bystander signals via extrinsic route and via Gap junctions. The model has been validated by experimental results in the literature. The time evolution of the effect and the dose-response curves were obtained in good accordance to them. Simulations were conducted for different values of bystander and irradiated cell densities with constant dose. From this work, we have obtained a relationship between cell density and effect. (author)

  9. Novel method for continuous cell separation by density gradient centrifugation: evaluation of a miniature separation column.

    Science.gov (United States)

    Shiono, Hiroyuki; Ito, Yoichiro

    2003-05-01

    A compact bench-top model of the centrifuge enables continuous cell separation based on density differences. The apparatus holds a small separation disk equipped with a circular channel (8 mL capacity) separated by a septum. A set of isotonic Percoll media with different densities is continuously introduced at one terminal and collected from the other. Under a centrifugal force field, cell suspension introduced into the proximal portion of the channel results in continuous separation of cells according to their densities. The performance of the apparatus was demonstrated with the separation of human buffy coat containing nucleated cells (>10(8)) among a large population (10(10)) of RBC. The results indicated that the method is capable of separating a large number of nucleated cells, with minimum damage, for a few hours of operation wherein neutrophils are well resolved from lymphocytes. The method may be applied to other types of samples including cord blood, blood from small animals, cultured cells, pancreatic beta cell islets, malaria parasites, sperm cells, etc.

  10. LTE Micro-cell Deployment for High-Density Railway Areas

    DEFF Research Database (Denmark)

    Sniady, Aleksander; Kassab, Mohamed; Soler, José

    2014-01-01

    Long Term Evolution (LTE) is a serious candidate for the future releases of the European Rail Traffic Management System (ERTMS). LTE offers more capacity and supports new communication-based applications and services for railways. Nevertheless, even with this technology, the classical macro......-cell radio deployments reach overload, especially in high-density areas, such as major train stations. In this paper, an LTE micro-cell deployment is investigated in high-density railway areas. Copenhagen Main Station is considered as a realistic deployment study case, with a set of relevant railway...

  11. Density of states measurements in a p-i-n solar cell

    Energy Technology Data Exchange (ETDEWEB)

    Crandall, R.S.; Wang, Q. [National Renewable Energy Lab., Golden, CO (United States)

    1996-05-01

    The authors describe results of density of states (DOS) profiling in p-i-n solar-cell devices using drive-level capacitance (DLC) techniques. Near the p-i interface the defect density is high, decreasing rapidly into the interior, reaching low values in the central region of the cell, and rising rapidly again at the n-i interface. They show that the states in the central region are neutral dangling-bond defects, whereas those near the interfaces with the doped layers are charged dangling bonds.

  12. The effects of daily wear contact lenses on goblet cell density.

    Science.gov (United States)

    Connor, C G; Campbell, J B; Steel, S A; Burke, J H

    1994-11-01

    Some patients can wear contact lenses with a low tear breakup time while others with an identical tear breakup time cannot wear lenses. This suggests the current method of tear film assessment is inadequate at differentiating between these two types of patients. The study attempts to expand our knowledge of the tear film with special attention directed to a critical yet little studied component: mucin. Mucin is vital to maintenance of the tear film and functions as a tear film stabilizer. The condition of the precorneal tear film is a major determinant in the success of contact lens wear. Eighteen subjects free of ocular surface disease who had never worn contact lenses had the goblet cell density of their inferior bulbar conjunctiva determined by impression cytology. The subjects were then fit in a 38 percent water polymacon lens and their goblet cell density determined on a monthly basis for 6 months. Nearly a 2-fold increase in goblet cell density was observed in 88 percent of the subjects over the 6-month period. The first statistically significant increase occurred 5 months after the initiation of lens wear when the goblet cell density rose from 4.19-7.84 percent. We speculate the increase in goblet cells is an adaptive response of the ocular surface to a coated daily wear contact lens.

  13. Limbal stem cell and oral mucosal epithelial transplantation from ex vivo cultivation in LSCD-induced rabbits: histology and immunologic study of the transplant epithelial sheet.

    Science.gov (United States)

    Tananuvat, Napaporn; Bumroongkit, Kanokkan; Tocharusa, Chainarong; Mevatee, Umnat; Kongkaew, Aphisek; Ausayakhun, Somsanguan

    2017-12-01

    To evaluate the results of cultivated limbal epithelial and oral mucosal epithelial transplantation (CLET and COMET) in limbal stem cell deficiency (LSCD)-induced rabbit model. Six New Zealand white rabbits were divided into two groups of three rabbits each. Limbal tissue was harvested from the first group, and oral mucosal biopsy was obtained from the second group. The tissues were cultured using an explant technique with amniotic membrane as a substrate and co-culture with the 3T3 fibroblast and air-lifting method. The right eye of each rabbit was induced to have LSCD using alkali burns. After three weeks, the LSCD-induced rabbit eyes were transplanted with the cultivated limbal and oral mucosal epithelial sheet in the first and second group, respectively. The transplanted eye was evaluated weekly post-operation. After 2 months, all transplanted eyes were enucleated and the epithelial morphology and phenotype of ocular surfaces were studied and compared with normal corneal and oral mucosal tissue. At 2-month post-transplantation, the eyes of four animals recovered with corneal transparency, one partially recovered, and one failed. The histology of the majority of transplanted eyes was stratified layers of corneal epithelia similar to normal rabbit cornea with some different findings such as goblet cells in the limbal region. Corneal epithelial thickening and stromal vascularization in two animals were observed. Phenotypic characterization of transplanted eyes showed a similar pattern of marker expression with the absence of p63 expression in the limbal or corneal epithelium in the COMET group. The histology and phenotype of transplanted eyes after CLET and COMET were most likely to have similar characteristics as a normal healthy rabbit eye even though the COMET eyes have some inferior characteristics to the CLET eyes.

  14. Spontaneous oscillations of cell voltage, power density, and anode exit CO concentration in a PEM fuel cell.

    Science.gov (United States)

    Lu, Hui; Rihko-Struckmann, Liisa; Sundmacher, Kai

    2011-10-28

    The spontaneous oscillations of the cell voltage and output power density of a PEMFC (with PtRu/C anode) using CO-containing H(2) streams as anodic fuels have been observed during galvanostatic operating. It is ascribed to the dynamic coupling of the CO adsorption (poisoning) and the electrochemical CO oxidation (reactivating) processes in the anode chamber of the single PEMFC. Accompanying the cell voltage and power density oscillations, the discrete CO concentration oscillations at the anode outlet of the PEMFC were also detected, which directly confirms the electrochemical CO oxidation taking place in the anode chamber during galvanostatic operating. This journal is © the Owner Societies 2011

  15. Respostas do feijão-vagem cultivado sob proteção com agrotêxtil em duas densidades de plantas Response of snap beans cultivated in two plant densities under nonwowen protection in Brazil

    Directory of Open Access Journals (Sweden)

    Adalberto Vitor Pereira

    2003-09-01

    yield of two cultivars of snap beans was evaluated. The current work was carried out at an experimental area of the Universidade Estadual de Ponta Grossa, Paraná State, Brazil. The experimental design was a completely randomized in split plots 2x2x2 (cultivars x protection x plant density, with four replications. The cultivars Turmalina and Coralina were sown in March (autumn, 2000, in two plant densities (6.6 and 3.3 pl m-1 under nonwowen protection (PP and natural environment (AN. Early yield, total yield, number and weight of pods were evaluated. The air (Tar and soil temperature amplitudes (Ts under PP and AN were also evaluated. Nonwoven protection and plant density did not influence the early yield. However, the use of nonwoven protection increased yield of 'Turmalina' as a result of higher number of pods per plant, compared with AN. There was no difference between PP and AN on yield of 'Coralina'. The plant density did not influence yield response of 'Turmalina'. On the other hand, 'Coralina' was more productive when cultivated under plant density of 3,3 pl m-1, when compared to 6,6 pl m-1. 'Coralina' was more productive than 'Turmalina' when cultivated under 3,3 pl m-1, however without differences between cultivars under 6,6 pl m-1. Air and soil temperature were higher under PP compared to AN, during the whole cycle of the culture, with medium differences between PP and AN of 2,63 and 1,97ºC, respectively for Tar and Ts. The difference between PP and AN, for Tar and Ts, was higher at the beginning of plant growing and decreased with the development of snap bean.

  16. Degradation of Solid Oxide Electrolysis Cells Operated at High Current Densities

    DEFF Research Database (Denmark)

    Tao, Youkun; Ebbesen, Sune Dalgaard; Mogensen, Mogens Bjerg

    2014-01-01

    In this work the durability of solid oxide cells for co-electrolysis of steam and carbon dioxide (45 % H2O + 45 % CO2 + 10 % H2) at high current densities was investigated. The tested cells are Ni-YSZ electrode supported, with a YSZ electrolyte and either a LSM-YSZ or LSCF-CGO oxygen electrode....... A current density of -1.5 and -2.0 A/cm2 was applied to the cell and the gas conversion was 45 % and 60 %, respectively. The cells were operated for a period of up to 700 hours. The electrochemical analysis revealed significant performance degradation for the ohmic process, oxygen ion interfacial transfer...

  17. A mathematical model of the current density distribution in electrochemical cells - AUTHORS’ REVIEW

    Directory of Open Access Journals (Sweden)

    PREDRAG M. ŽIVKOVIĆ

    2011-06-01

    Full Text Available An approach based on the equations of electrochemical kinetics for the estimation of the current density distribution in electrochemical cells is presented. This approach was employed for a theoretical explanation of the phenomena of the edge and corner effects. The effects of the geometry of the system, the kinetic parameters of the cathode reactions and the resistivity of the solution are also discussed. A procedure for a complete analysis of the current distribution in electrochemical cells is presented.

  18. Inverse Relationship Between Leydig Cell Density and Metastatic Potential of Prostatic Adenocarcinoma

    Directory of Open Access Journals (Sweden)

    W. John Wang

    1999-01-01

    Full Text Available Purpose: Evaluate the relationship between metastatic potential of prostatic adenocarcinoma (PC and testicular Leydig cell density. Materials and methods: Tissue samples from 111 men, age 52–85, with PC and bilateral orchiectomy were evaluated for Leydig cell density. The patients were divided into two groups: Group A were patients with metastasis (n=36 and Group B were patients without metastasis (n=75. Leydig cell density was determined by direct manual microscopic cell count on the tissue sections. The means of cell counts by four pathologists, expressed as cell/0.78 mm2 were used for analysis. The normally distributed data were analyzed by two‐tail Student’s t‐test. Thirty‐eight age‐compatible autopsy cases who died of unrelated causes served as normal controls. Results: The mean of Leydig cell count in group A patients was 14.43 (14.43 ± 1.19 SE. Mean of Group B was 47.05 (47.05 ± 4.05 SE whereas normal controls displayed a mean of 48.66 (48.66 ± 2.94 SE. Group A was significantly different from control (p0.75. Conclusions: Patients with metastatic adenocarcinoma of prostate, as a group, have a significantly lower Leydig cell density than patients without metastasis or patients without PC in compatible age groups. The hormonal relationship between this observation is however unknown. One possible explanation is that PC subpopulation with metastatic potential may require different level of endogenous androgen or are androgen‐independent.

  19. Antcin K, an active triterpenoid from the fruiting bodies of basswood cultivated Antrodia cinnamomea, induces mitochondria and endoplasmic reticulum stress-mediated apoptosis in human hepatoma cells

    Directory of Open Access Journals (Sweden)

    Chiao-I. Lai

    2016-01-01

    Full Text Available Liver cancer is the second leading cause of cancer deaths in Taiwan as per the 2011 statistics and ranks fourth in cancer-related mortality in the world. Recent researches have shown that Antrodia cinnamomea, a Taiwan-specific medicinal mushroom, has biological activities, including hepatoprotection, anti-inflammation, antihepatitis B virus activity, and anticancer activity. In the present study, the antiproliferative activity and molecular mechanisms of antcin K, the most abundant ergostane triterpenoid from the fruiting bodies of basswood cultivated A. cinnamomea, were investigated using human hepatoma Hep 3B cells. The results showed that antcin K effectively reduced Hep 3B cells viability within 48 hours. Antcin K induced phosphatidylserine exposure, chromatin condensation, and DNA damage, but did not significantly increase autophagosome content or cause cell expansion and cell lysis. Thus, the principal mode of Hep 3B cells death induced by antcin K was apoptosis, rather than autophagy or necrosis. In-depth investigation of the molecular mechanisms revealed that antcin K first promoted reactive oxygen species generation and adenosine triphosphate depletion, leading to endoplasmic reticulum stress and resulting in mitochondrial membrane permeability changes. After losing the mitochondrial membrane potential, caspase-independent and caspase-dependent apoptosis-related proteins were released, including HtrA2, apoptotic-induced factor, endonuclease G, and cytochrome c. Cytochrome c activated caspase-9 and caspase-3, and cut downstream protein PARP, ultimately leading to cell apoptosis. These results suggested that antcin K induced mitochondrial and endoplasmic reticulum stress-mediated apoptosis in human hepatoma cells. Coupled with these findings, antcin K has a potential to be a complementary agent in liver cancer therapy.

  20. Absolute choline concentration measured by quantitative proton MR spectroscopy correlates with cell density in meningioma

    Energy Technology Data Exchange (ETDEWEB)

    Yue, Qiang [University of Tsukuba, Department of Neurosurgery, Institute of Clinical Medicine, Tsukuba Science City, Ibaraki (Japan)]|[West China Hospital of Sichuan University, Huaxi MR Research Center, Department of Radiology, Chengdu (China); Shibata, Yasushi; Kawamura, Hiraku; Matsumura, Akira [University of Tsukuba, Department of Neurosurgery, Institute of Clinical Medicine, Tsukuba Science City, Ibaraki (Japan); Isobe, Tomonori [Kitasato University, Department of Medical Technology, School of Allied Health Sciences, Minato, Tokyo (Japan); Anno, Izumi [University of Tsukuba, Department of Radiology, Institute of Clinical Medicine, Tsukuba, Ibaraki (Japan); Gong, Qi-Yong [West China Hospital of Sichuan University, Huaxi MR Research Center, Department of Radiology, Chengdu (China)]|[University of Liverpool, Division of Medical Imaging, Faculty of Medicine, Liverpool (United Kingdom)

    2009-01-15

    This study was aimed to investigate the relationship between quantitative proton magnetic resonance spectroscopy (1H-MRS) and pathological changes in meningioma. Twenty-two meningioma cases underwent single voxel 1H-MRS (point-resolved spectroscopy sequence, repetition time/echo time = 2,000 ms/68, 136, 272 ms). Absolute choline (Cho) concentration was calculated using tissue water as the internal reference and corrected according to intra-voxel cystic/necrotic parts. Pathological specimens were stained with MIB-1 antibody to measure cell density and proliferation index. Correlation analysis was performed between absolute Cho concentration and cell density and MIB-1 labeled proliferation index. Average Cho concentration of all meningiomas before correction was 2.95 {+-} 0.86 mmol/kg wet weight. It was increased to 3.23 {+-} 1.15 mmol/kg wet weight after correction. Average cell density of all meningiomas was 333 {+-} 119 cells/HPF, and average proliferation index was 2.93 {+-} 5.72%. A linear, positive correlation between cell density and Cho concentration was observed (r = 0.650, P = 0.001). After correction of Cho concentration, the correlation became more significant (r = 0.737, P < 0.001). However, no significant correlation between Cho concentration and proliferation index was found. There seemed to be a positive correlation trend after correction of Cho concentration but did not reach significant level. Absolute Cho concentration, especially Cho concentration corrected according to intra-voxel cystic/necrotic parts, reflects cell density of meningioma. (orig.)

  1. Disabled cell density sensing leads to dysregulated cholesterol synthesis in glioblastoma.

    Science.gov (United States)

    Kambach, Diane M; Halim, Alan S; Cauer, A Gesine; Sun, Qian; Tristan, Carlos A; Celiku, Orieta; Kesarwala, Aparna H; Shankavaram, Uma; Batchelor, Eric; Stommel, Jayne M

    2017-02-28

    A hallmark of cellular transformation is the evasion of contact-dependent inhibition of growth. To find new therapeutic targets for glioblastoma, we looked for pathways that are inhibited by high cell density in astrocytes but not in glioma cells. Here we report that glioma cells have disabled the normal controls on cholesterol synthesis. At high cell density, astrocytes turn off cholesterol synthesis genes and have low cholesterol levels, but glioma cells keep this pathway on and maintain high cholesterol. Correspondingly, cholesterol pathway upregulation is associated with poor prognosis in glioblastoma patients. Densely-plated glioma cells increase oxygen consumption, aerobic glycolysis, and the pentose phosphate pathway to synthesize cholesterol, resulting in a decrease in reactive oxygen species, TCA cycle intermediates, and ATP. This constitutive cholesterol synthesis is controlled by the cell cycle, as it can be turned off by cyclin-dependent kinase inhibitors and it correlates with disabled cell cycle control though loss of p53 and RB. Finally, glioma cells, but not astrocytes, are sensitive to cholesterol synthesis inhibition downstream of the mevalonate pathway, suggesting that specifically targeting cholesterol synthesis might be an effective treatment for glioblastoma.

  2. Ultra-high cell-density silicon photomultipliers with high detection efficiency

    Science.gov (United States)

    Acerbi, Fabio; Gola, Alberto; Regazzoni, Veronica; Paternoster, Giovanni; Borghi, Giacomo; Piemonte, Claudio; Zorzi, Nicola

    2017-05-01

    Silicon photomultipliers (SiPMs) are arrays of many single-photon avalanche diodes (SPADs), all connected in parallel. Each SPAD is sensitive to single photons and the SiPM gives an output proportional to the number of detected photons. These sensors are becoming more and more popular in different applications, from high-energy physics to spectroscopy, and they have been significantly improved over last years, decreasing the noise, increasing the cell fill-factor (FF) and thus achieving very high photon-detection efficiency (PDE). In FBK (Trento, Italy), we developed new SiPM technologies with high-density (HD) and, more recently, ultra-high-density (UHD) of cells (i.e. density of SPADs). These technologies employ deep-trenches between cells, for electrical and optical isolation. As an extreme case the smallest-cell, SiPM, i.e. with 5μm cell pitch, has about 40000 SPADs per squared millimeter. Such small SPAD dimensions gives a significantly high dynamic range to the SiPM. These small-cells SiPM have a lower correlated noise (including lower afterpulsing probability) and a faster recharge time (in the order of few nanoseconds), and they also preserve a very good detection efficiency (despite the small SPAD dimension).

  3. Cell openness manipulation of low density polyurethane foam for efficient sound absorption

    Science.gov (United States)

    Hyuk Park, Ju; Suh Minn, Kyung; Rae Lee, Hyeong; Hyun Yang, Sei; Bin Yu, Cheng; Yeol Pak, Seong; Sung Oh, Chi; Seok Song, Young; June Kang, Yeon; Ryoun Youn, Jae

    2017-10-01

    Satisfactory sound absorption using a low mass density foam is an intriguing desire for achieving high fuel efficiency of vehicles. This issue has been dealt with a microcellular geometry manipulation. In this study, we demonstrate the relationship between cell openness of polyurethane (PU) foam and sound absorption behaviors, both theoretically and experimentally. The objective of this work is to mitigate a threshold of mass density by rendering a sound absorber which shows a satisfactory performance. The cell openness, which causes the best sound absorption performance in all cases considered, was estimated as 15% by numerical simulation. Cell openness of PU foam was experimentally manipulated into desired ranges by adjusting rheological properties in a foaming reaction. Microcellular structures of the fabricated PU foams were observed and sound absorption coefficients were measured using a B&K impedance tube. The fabricated PU foam with the best cell openness showed better sound absorption performance than the foam with double mass density. We envisage that this study can help the manufacture of low mass density sound absorbing foams more efficiently and economically.

  4. Corneal endothelial cell density and morphology in low and moderate myopic Chinese eyes

    Directory of Open Access Journals (Sweden)

    Jane Mei Chun

    2013-08-01

    Full Text Available AIM: To describe and compare the corneal endothelial cell density and morphology in young, low and moderate myopic Chinese adults in Malaysian Chinese population.METHODS: Non-contact specular microscopy (Topcon SP3000P, Tokyo, Japan was performed in low (n=78; 21.22±1.51 years and moderate (n=78; 21.82±1.40 years myopic subjects. The mean of three consecutive measurements of endothelial cell density (MCD, coefficient of variation (CV in the cell size, and hexagonal appearance of the cell were obtained.RESULTS: In low myopic eyes the MCD was 3 063.0±176.2/mm2, the mean CV was 33.4±4.0% and the mean hexagonal appearance of the cell was 57.9±2.7%. In moderate myopic eyes the MCD was 2961.6±159.0/mm2, the mean CV was 33.9±3.6% and mean hexagonal appearance of the cell was 56.2±4.7%. There were statistically significant differences in MCD (PPCONCLUSION:The corneal endothelial cell layer in more myopic eyes tends to have less MCD and cell hexagonality compared to lower myopic eyes. Nevertheless, there is no significant difference in CV between low and moderate myopic eyes.

  5. Influence of Oxygen in the Cultivation of Human Mesenchymal Stem Cells in Simulated Microgravity: An Explorative Study

    NARCIS (Netherlands)

    Versari, S.; Klein-Nulend, J.; van Loon, J.J.W.A.; Bradamante, S.

    2013-01-01

    Previous studies indicated that human Adipose Tissue-derived Mesenchymal Stem Cells (AT-MSCs) cultured in simulated microgravity (sim-μg) in standard laboratory incubators alter their proliferation and differentiation. Recent studies on the stem cell (SC) niches and the influence of oxygen on SC

  6. Influence of oxygen in the cultivation of human mesenchymal stem cells in simulated microgravity: an explorative study

    NARCIS (Netherlands)

    Versari, S.; Klein-Nulend, J.; van Loon, J.; Bradamante, S.

    2013-01-01

    Previous studies indicated that human Adipose Tissue-derived Mesenchymal Stem Cells (AT-MSCs) cultured in simulated microgravity (sim-μg) in standard laboratory incubators alter their proliferation and differentiation. Recent studies on the stem cell (SC) niches and the influence of oxygen on SC

  7. A figure of merit to evaluate transparent conductor oxides for solar cells using photonic flux density

    Energy Technology Data Exchange (ETDEWEB)

    Mendez-Gamboa, J.A., E-mail: jmendez@uady.mx [Engineering School, University of Yucatan, AP 150 Cordemex, 97310 Mérida, Yucatán (Mexico); Castro-Rodriguez, R. [Applied Physics Department, CINVESTAV-IPN Mérida, C.P. 97310, Mérida, Yucatán (Mexico); Perez-Quintana, I.V.; Medina-Esquivel, R.A.; Martel-Arbelo, A. [Engineering School, University of Yucatan, AP 150 Cordemex, 97310 Mérida, Yucatán (Mexico)

    2016-01-29

    We report an alternative method to evaluate transparent conductor oxides (TCO) from their photonic flux density (PFD(hυ)) to be used in solar cells. From the transmittance spectrum (T(hυ)) in the visible region, we calculate the PFD(hυ) and the solar photon flux-weighted transmittance (T{sub SW}) of one specific TCO with potential application in solar cells. The photo-current density (J{sub PH}) in mA/cm{sup 2} of one specific TCO when exposed to white light is evaluated when PFD(hυ) is integrated over the whole solar electromagnetic spectrum. Finally, we define a figure of merit as J{sub PH} over the TCO film sheet resistance to find the best equilibrium between the transmission and its electrical resistance. To carry out this work, a bibliographical search of investigations about development of TCOs was extensively made to evaluate its T(hυ), T{sub SW,} PFD(hυ), J{sub PH} and the figure of merit that we propose. From our results, we consider that the proposed method is a good tool for a fine comparison of transparent conductive films in solar cell development. - Highlights: • A figure of merit is presented for the evaluation of TCO used in solar cells. • It is defined as the ratio of photo-current density over TCO sheet resistance. • It is a tool for a fine comparison of TCO films in solar cell development.

  8. Isolation and cultivation of fungal strains from in vitro cell cultures of two marine sponges (Porifera: Halichondrida and Haplosclerida

    Directory of Open Access Journals (Sweden)

    Enrique E. Rozas

    2011-12-01

    Full Text Available Despite the large number of reports describing sponge-microbe associations, limited knowledge is available about associated fungi and their relationships with the hosts. In this work, specific fungal strains were obtained directly from in vitro sponge cell cultures (primmorphs and single sponge cells (cytospins and compared with those obtained from whole tissue preparations. A total of 27 fungal strains were isolated from the marine sponges Hymeniacidon heliophila and Haliclona melana. Fifteen strains, nine from H. heliophila and six from H. melana, were obtained from whole tissue and were considered as possible mesohyl associated or transient fungi. Twelve strains were isolated from in vitro sponge cell cultures (primmorphs and were, therefore, considered as cell associated. From these, five different strains were obtained from H. heliophila isolated cells, while five were identified from cytospins and two from primmorphs of H. melana. The fungal strains obtained from cell cultures from both sponge species were different, and none of them were detected in the whole tissue preparations of the same species. Nine H. heliophila and seven H. melana strains shows low similarity with the sequences available in public databases and belong to potentially new species. This is the first report of fungi isolated directly from sponge cells, which allowed the observation and selection of specific strains that probably would not be obtained by usual culture dependent techniques.

  9. Noninvasive prenatal diagnosis. Use of density gradient centrifugation, magnetically activated cell sorting and in situ hybridization

    DEFF Research Database (Denmark)

    Campagnoli, C; Multhaupt, H A; Ludomirski, A

    1997-01-01

    cells recovered did not differ. Seven of seven male pregnancies were correctly identified. One case of trisomy 21 was detected. CONCLUSION: The in situ hybridization analysis of fetal nucleated erythrocytes isolated from maternal blood using single density gradient centrifugation, anti-CD71/anti...... of the isolated cells were subjected to in situ hybridization with specific DNA probes for the Y chromosome and chromosome 21 to confirm the fetal origin. RESULTS: After MiniMACS the enrichment factors for the CD71/GPA- and CD36/GPA-positive cells from maternal blood were similar, and the percentages of fetal...

  10. In vitro culture of cell aggregates of rye (Secale cereale L.) at low densities.

    Science.gov (United States)

    Mottley, J; Sybenga, J

    1988-05-01

    The growth of cell aggregates from a rye suspension culture was tested at low density in three culture systems. Mass seeding was the most supportive system, followed by agarose droplets. Microdroplet culture using Cuprak dishes was the least effective system. Growth was stimulated by the presence of a feeder layer of suspension cells but only if the feeder contact with the nursed cells was via a liquid and not a gaseous phase. Plating efficiences were enhanced by the feeder effect, whereas the subsequent growth rates were less affected. The techniques described should prove useful in programs aimed at the in vitro genetic manipulation of rye or other cereals.

  11. A High Power-Density Mediator-Free Microfluidic Biophotovoltaic Device for Cyanobacterial Cells

    CERN Document Server

    Bombelli, Paolo; Herling, Therese W; Howe, Christopher J; Knowles, Tuomas P J

    2014-01-01

    Biophotovoltaics has emerged as a promising technology for generating renewable energy since it relies on living organisms as inexpensive, self-repairing and readily available catalysts to produce electricity from an abundant resource - sunlight. The efficiency of biophotovoltaic cells, however, has remained significantly lower than that achievable through synthetic materials. Here, we devise a platform to harness the large power densities afforded by miniaturised geometries. To this effect, we have developed a soft-lithography approach for the fabrication of microfluidic biophotovoltaic devices that do not require membranes or mediators. Synechocystis sp. PCC 6803 cells were injected and allowed to settle on the anode, permitting the physical proximity between cells and electrode required for mediator-free operation. We demonstrate power densities of above 100 mW/m2 for a chlorophyll concentration of 100 {\\mu}M under white light, a high value for biophotovoltaic devices without extrinsic supply of additional...

  12. Renaturation and purification of bone morphogenetic protein-2 produced as inclusion bodies in high-cell-density cultures of recombinant Escherichia coli.

    Science.gov (United States)

    Vallejo, Luis Felipe; Brokelmann, Maren; Marten, Sabine; Trappe, Susanne; Cabrera-Crespo, Joaquin; Hoffmann, Andrea; Gross, Gerhard; Weich, Herbert A; Rinas, Ursula

    2002-03-28

    Eschericha coli was genetically engineered to produce recombinant human bone morphogenetic protein-2 (rhBMP-2) in a non-active aggregated form using a temperature-inducible expression system. High concentrations of both biomass (75 g cell dry weight per liter of culture broth) and inactive rhBMP-2 (8.6 gl(-1)) were obtained by applying a high-cell-density cultivation procedure. After washing and solubilizing the inclusion bodies, rhBMP-2 was refolded and dimerized at concentrations up to 100 mgl(-1) by means of a simple dilution method with yields exceeding 50%. Finally, a one-step purification procedure based on affinity chromatography was implemented to isolate the rhBMP-2 dimer. With the established renaturation and purification protocols, yields of more than 10 mg rhBMP-2 dimer per gram cell dry weight were obtained corresponding to 750 mg rhBMP-2 dimer per liter of culture broth. The purified rhBMP-2 dimer showed biological activity equivalent to CHO produced rhBMP-2 as tested by the induction of alkaline phosphatase activity in C2C12 cells.

  13. The role of oxygen in yeast metabolism during high cell density brewery fermentations.

    Science.gov (United States)

    Verbelen, P J; Saerens, S M G; Van Mulders, S E; Delvaux, F; Delvaux, F R

    2009-04-01

    The volumetric productivity of the beer fermentation process can be increased by using a higher pitching rate (i.e., higher inoculum size). However, the decreased yeast net growth observed in these high cell density fermentations can have a negative impact on the physiological stability throughout subsequent yeast generations. The use of different oxygen conditions (wort aeration, wort oxygenation, yeast preoxygenation) was investigated to improve the growth yield during high cell density fermentations and yeast metabolic and physiological parameters were assessed systematically. Together with a higher extent of growth (dependent on the applied oxygen conditions), the fermentation power and the formation of unsaturated fatty acids were also affected. Wort oxygenation had a significant decreasing effect on the formation of esters, which was caused by a decreased expression of the alcohol acetyl transferase gene ATF1, compared with the other conditions. Lower glycogen and trehalose levels at the end of fermentation were observed in case of the high cell density fermentations with oxygenated wort and the reference fermentation. The expression levels of BAP2 (encoding the branched chain amino acid permease), ERG1 (encoding squalene epoxidase), and the stress responsive gene HSP12 were predominantly influenced by the high cell concentrations, while OLE1 (encoding the fatty acid desaturase) and the oxidative stress responsive genes SOD1 and CTT1 were mainly affected by the oxygen availability per cell. These results demonstrate that optimisation of high cell density fermentations could be achieved by improving the oxygen conditions, without drastically affecting the physiological condition of the yeast and beer quality.

  14. A high parasite density environment induces transcriptional changes and cell death in Plasmodium falciparum blood stages.

    Science.gov (United States)

    Chou, Evelyn S; Abidi, Sabia Z; Teye, Marian; Leliwa-Sytek, Aleksandra; Rask, Thomas S; Cobbold, Simon A; Tonkin-Hill, Gerry Q; Subramaniam, Krishanthi S; Sexton, Anna E; Creek, Darren J; Daily, Johanna P; Duffy, Michael F; Day, Karen P

    2017-12-27

    Transient regulation of Plasmodium numbers below the density that induces fever has been observed in chronic malaria infections in humans. This species transcending control cannot be explained by immunity alone. Using an in vitro system we have observed density dependent regulation of malaria population size as a mechanism to possibly explain these in vivo observations. Specifically, Plasmodium falciparum blood stages from a high but not low-density environment exhibited unique phenotypic changes during the late trophozoite (LT) and schizont stages of the intraerythrocytic cycle. These included in order of appearance: failure of schizonts to mature and merozoites to replicate, apoptotic-like morphological changes including shrinking, loss of mitochondrial membrane potential, and blebbing with eventual release of aberrant parasites from infected erythrocytes. This unique death phenotype was triggered in a stage-specific manner by sensing of a high-density culture environment. Conditions of glucose starvation, nutrient depletion, and high lactate could not induce the phenotype. A high-density culture environment induced rapid global changes in the parasite transcriptome including differential expression of genes involved in cell remodeling, clonal antigenic variation, metabolism, and cell death pathways including an apoptosis-associated metacaspase gene. This transcriptional profile was also characterized by concomitant expression of asexual and sexual stage-specific genes. The data show strong evidence to support our hypothesis that density sensing exists in P. falciparum. They indicate that an apoptotic-like mechanism may play a role in P. falciparum density regulation, which, as in yeast, has features quite distinguishable from mammalian apoptosis. Gene expression data are available in the GEO databases under the accession number GSE91188. © 2017 Federation of European Biochemical Societies.

  15. Anatomical Mapping and Density of Merkel Cells in Skin and Mucosae of the Dog.

    Science.gov (United States)

    Ramírez, Gustavo A; Rodríguez, Francisco; Quesada, Óscar; Herráez, Pedro; Fernández, Antonio; Espinosa-de-Los-Monteros, Antonio

    2016-09-01

    Merkel cells (MCs) are specialized cutaneous receptor cells involved with tactile sense. Although the distribution of MCs has been extensively studied in humans and rodents, their precise distribution and density throughout skin in the dog has not previously been determined. Knowledge of their distribution could facilitate understanding of their functions. By using of immunohistochemistry, density, and anatomical mapping of the MCs population in the dog skin was determined. Assessment of the MCs innervation was also achieved. Different patterns were noted in epidermis, hair follicles, or mucosa, including variable-sized clusters, linear or horse-shaped arrangements, and scattered and individualized cells. MCs revealed great variations in density and distribution over the body surface, with the highest numbers in oral mucosa and facial skin. There was no correlation of MCs density with age, sex, type of breed, coat type or pigmentation. Between 41 and 65% of MCs in hairy and glabrous skin and 8-18% of MCs in oral mucosa were in intimate contact with intraepithelial axon terminals. These findings indicate that canine MCs are numerous in sensory receptive areas and may be associated with the tactile sense in the dog. The present article enhances the knowledge of the skin structure in this species. Anat Rec, 299:1157-1164, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Spirulina cultivation in China

    Science.gov (United States)

    Wu, Bo-Tang; Xiang, Wen-Zhou; Zeng, Cheng-Kui

    1998-03-01

    This paper reviews and discusses the development and many problems of Spirulina cultivation in China, points out the advantages and disadvantages of open photobioreactor system, and predicts that seawater Spirulina cultivation will be a new trend to be strengthened and emphasized due to its special physiological characteristics, easier management, lower fertilizer cost, and higher resistance to contaminants and rare pollution of chemicals.

  17. Direct alcohol fuel cells: toward the power densities of hydrogen-fed proton exchange membrane fuel cells.

    Science.gov (United States)

    Chen, Yanxin; Bellini, Marco; Bevilacqua, Manuela; Fornasiero, Paolo; Lavacchi, Alessandro; Miller, Hamish A; Wang, Lianqin; Vizza, Francesco

    2015-02-01

    A 2 μm thick layer of TiO2 nanotube arrays was prepared on the surface of the Ti fibers of a nonwoven web electrode. After it was doped with Pd nanoparticles (1.5 mgPd  cm(-2) ), this anode was employed in a direct alcohol fuel cell. Peak power densities of 210, 170, and 160 mW cm(-2) at 80 °C were produced if the cell was fed with 10 wt % aqueous solutions of ethanol, ethylene glycol, and glycerol, respectively, in 2 M aqueous KOH. The Pd loading of the anode was increased to 6 mg cm(-2) by combining four single electrodes to produce a maximum peak power density with ethanol at 80 °C of 335 mW cm(-2) . Such high power densities result from a combination of the open 3 D structure of the anode electrode and the high electrochemically active surface area of the Pd catalyst, which promote very fast kinetics for alcohol electro-oxidation. The peak power and current densities obtained with ethanol at 80 °C approach the output of H2 -fed proton exchange membrane fuel cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Mathematical model of a rotational bioreactor for the dynamic cultivation of scaffold-adhered human mesenchymal stem cells for bone regeneration

    Science.gov (United States)

    Ganimedov, V. L.; Papaeva, E. O.; Maslov, N. A.; Larionov, P. M.

    2017-09-01

    Development of cell-mediated scaffold technologies for the treatment of critical bone defects is very important for the purpose of reparative bone regeneration. Today the properties of the bioreactor for cell-seeded scaffold cultivation are the subject of intensive research. We used the mathematical modeling of rotational reactor and construct computational algorithm with the help of ANSYS software package to develop this new procedure. The solution obtained with the help of the constructed computational algorithm is in good agreement with the analytical solution of Couette for the task of two coaxial cylinders. The series of flow computations for different rotation frequencies (1, 0.75, 0.5, 0.33, 1.125 Hz) was performed for the laminar flow regime approximation with the help of computational algorithm. It was found that Taylor vortices appear in the annular gap between the cylinders in a simulated bioreactor. It was obtained that shear stress in the range of interest (0.002-0.1 Pa) arise on outer surface of inner cylinder when it rotates with the frequency not exceeding 0.8 Hz. So the constructed mathematical model and the created computational algorithm for calculating the flow parameters allow predicting the shear stress and pressure values depending on the rotation frequency and geometric parameters, as well as optimizing the operating mode of the bioreactor.

  19. A device to facilitate preparation of high-density neural cell cultures in MEAs.

    Science.gov (United States)

    Mok, S Y; Lim, Y M; Goh, S Y

    2009-05-15

    A device to facilitate high-density seeding of dissociated neural cells on planar multi-electrode arrays (MEAs) is presented in this paper. The device comprises a metal cover with two concentric cylinders-the outer cylinder fits tightly on to the external diameter of a MEA to hold it in place and an inner cylinder holds a central glass tube for introducing a cell suspension over the electrode area of the MEA. An O-ring is placed at the bottom of the inner cylinder and the glass tube to provide a fluid-tight seal between the glass tube and the MEA electrode surface. The volume of cell suspension in the glass tube is varied according to the desired plating density. After plating, the device can be lifted from the MEA without leaving any residue on the contact surface. The device has enabled us to increase and control the plating density of neural cell suspension with low viability, and to prepare successful primary cultures from cryopreserved neurons and glia. The cultures of cryopreserved dissociated cortical neurons that we have grown in this manner remained spontaneously active over months, exhibited stable development and similar network characteristics as reported by other researchers.

  20. Modeling Bubble Flow and Current Density Distribution in an Alkaline Electrolysis Cell

    Directory of Open Access Journals (Sweden)

    Ravichandra S. Jupudi

    2009-12-01

    Full Text Available The effect of bubbles on the current density distribution over the electrodes of an alkaline electrolyzer cell is studied using a two-dimensional computational fluid dynamics model. Model includes Eulerian-Eulerian two-phase flow methodology to model the multiphase flow of Hydrogen and Oxygen with water and the behavior of each phase is accounted for using first principle. Hydrogen/Oxygen evolution, flow field and current density distribution are incorporated in the model to account for the complicated physics involved in the process. Fluent 6.2 is used to solve two-phase flow and electrochemistry is incorporated using UDF (User Defined Function feature of Fluent. Model is validated with mesh refinement study and by comparison with experimental measurements. Model is found to replicate the effect of cell voltage and inter-electrode gap (distance between the electrodes on current density accurately. Further, model is found to capture the existence of optimum cell height. The validated model is expected to be a very useful tool in the design and optimization of alkaline electrolyzer cells.

  1. The invasive microalga Chrysophaeum taylorii: Interactive stressors regulate cell density and mucilage production.

    Science.gov (United States)

    Caronni, Sarah; Calabretti, Chiara; Cavagna, Gianluca; Ceccherelli, Giulia; Delaria, Maria Anna; Macri, Giovanni; Navone, Augusto; Panzalis, Pieraugusto

    2017-08-01

    The benthic mucilage producing microalga Chrysophaeum taylorii Lewis and Bryan (Pelagophyceae) has recently received attention for its rapid spread in the Mediterranean Sea, where its blooms have remarkable detrimental effects. So far no information on C. taylorii response to multiple stressors, especially in terms of mucilage hyperproduction, is available in the literature yet, and a manipulative field experiment in this topic was designed in Tavolara Punta Coda Cavallo Marine Protected Area. The aim of the study was to test the effects of nutrient enrichment (addition of nutrients), mechanical disturbance (partial and total benthic organisms removal) and hydrodynamics (increased water turbulence) on C. taylorii cell density and mucilage abundance. To the purpose, the three above mentioned stressors were simulated and the three treatments were assigned to 20 × 20 cm plots following a full-factorial design (n = 3). Interactive effects of the three stressors affected significantly both benthic C. taylorii cell density and mucilage cover although differently. Mechanical disturbance and high hydrodynamics produced consistent effects on cell density and mucilage production (i.e. the former factor enhancing and the latter decreasing). Nutrient enrichment on the contrary led to contrasting effects, promoting cell abundance and inhibiting mucilage production. Therefore, important mucilage blooms are expected in oligotrophic sheltered coastal locations where barren areas are present. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Poly(N-isopropylacrylamide) modified polydopamine as a temperature-responsive surface for cultivation and harvest of mesenchymal stem cells.

    Science.gov (United States)

    Zhang, Jun; Peng, Ching-An

    2017-10-24

    A thermo-responsive surface was fabricated by depositing poly(N-isopropylacrylamide) (PNIPAAm) onto polydopamine coated cell culture substrata through free radical polymerization for the purpose of culturing and harvesting human mesenchymal stem cells (hMSCs). Human MSCs were cultured onto the PNIPAAm-g-polydopamine coated surface and harvested by changing from physiological to ambient temperature. The produced PNIPAAm-g-polydopamine surface was characterized by atomic force microscopy, Fourier transform infrared spectroscopy, nuclear magnetic resonance, water contact angle measurement, differential scanning calorimetry, and cell culture studies. Our results revealed that hMSCs could be detached from the PNIPAAm-g-polydopamine surface within 60 min after switching the temperature from 37 °C to room temperature. The detached hMSCs were able to proliferate on the PNIPAAm-g-polydopamine coated surface for further growth and harvest.

  3. Increased cell proliferation and mucocyte density in the sea anemone Aiptasia pallida recovering from bleaching.

    Science.gov (United States)

    Fransolet, David; Roberty, Stéphane; Herman, Anne-Catherine; Tonk, Linda; Hoegh-Guldberg, Ove; Plumier, Jean-Christophe

    2013-01-01

    Recovery of coral after bleaching episodes is a critical period for the health of the reef ecosystem. While events such as symbiont (genus Symbiodinium) shifting/shuffling or tissue apoptosis have been demonstrated to occur following bleaching, little is known concerning tissue recovery or cell proliferation. Here, we studied the sea anemone Aiptasia pallida exposed to a transient elevation of water temperature combined with high illumination (33°C and 1900 µmol photons x m(-2) x s(-1) for 30 h). Following such treatment bleached anemones showed a significant reduction of their Symbiodinium density. Cell proliferation in the ectodermis and gastrodermis was determined by assessing the densities of cells labeled with a thymidine analogue (EdU). Cell proliferation significantly increased during the first day following stress in both tissue types. This increased cell proliferation returned to pre-stress values after one week. Although cell proliferation was higher in the ectodermis in absence of stress, it was relatively more pronounced in the gastrodermis of stressed anemones. In addition, the ratio of ectodermal mucocytes significantly increased three weeks after induced stress. These results suggest that thermal/photic stress coupled with the loss of the symbionts is able to enhance cell proliferation in both gastrodermis and ectodermis of cnidarians. While new cells formed in the gastrodermis are likely to host new Symbiodinium, the fate of new cells in the ectodermis was only partially revealed. Some new ectodermal cells may, in part, contribute to the increased number of mucocytes which could eventually help strengthen the heterotrophic state until restoration of the symbiosis.

  4. Increased cell proliferation and mucocyte density in the sea anemone Aiptasia pallida recovering from bleaching.

    Directory of Open Access Journals (Sweden)

    David Fransolet

    Full Text Available Recovery of coral after bleaching episodes is a critical period for the health of the reef ecosystem. While events such as symbiont (genus Symbiodinium shifting/shuffling or tissue apoptosis have been demonstrated to occur following bleaching, little is known concerning tissue recovery or cell proliferation. Here, we studied the sea anemone Aiptasia pallida exposed to a transient elevation of water temperature combined with high illumination (33°C and 1900 µmol photons x m(-2 x s(-1 for 30 h. Following such treatment bleached anemones showed a significant reduction of their Symbiodinium density. Cell proliferation in the ectodermis and gastrodermis was determined by assessing the densities of cells labeled with a thymidine analogue (EdU. Cell proliferation significantly increased during the first day following stress in both tissue types. This increased cell proliferation returned to pre-stress values after one week. Although cell proliferation was higher in the ectodermis in absence of stress, it was relatively more pronounced in the gastrodermis of stressed anemones. In addition, the ratio of ectodermal mucocytes significantly increased three weeks after induced stress. These results suggest that thermal/photic stress coupled with the loss of the symbionts is able to enhance cell proliferation in both gastrodermis and ectodermis of cnidarians. While new cells formed in the gastrodermis are likely to host new Symbiodinium, the fate of new cells in the ectodermis was only partially revealed. Some new ectodermal cells may, in part, contribute to the increased number of mucocytes which could eventually help strengthen the heterotrophic state until restoration of the symbiosis.

  5. Poda apical, densidade de plantas e cobertura plástica do solo na produtividade do tomateiro em cultivo protegido Tip pruning, plant density and plastic mulching in tomato yield in protected cultivation

    Directory of Open Access Journals (Sweden)

    Júlio César Bogiani

    2008-01-01

    Full Text Available O objetivo deste trabalho foi o de avaliar o efeito de duas combinações entre densidade de plantas e altura de poda apical, associados à utilização de cobertura plástica do solo, sobre o desenvolvimento e a produção de frutos do tomateiro, híbrido Duradoro (EMBRAPA, em ambiente protegido. Foram estabelecidos dois sistemas de cultivo com haste única: SC1 - poda apical após a 5ª inflorescência, com 20.000 plantas ha-1 e SC2 - poda apical após a 2ª inflorescência com 50.000 plantas ha-1; e três condições de cobertura do solo: solo nu; filme de polietileno branco e filme de polietileno preto, adotando-se o delineamento em blocos ao acaso, em esquema de parcelas subdivididas. O experimento foi desenvolvido de junho a novembro de 2003, na UNESP, Câmpus de Ilha Solteira. A adoção do tratamento SC2 determinou maior altura de plantas aos 45 dias após o transplante, maior número de frutos por área e maior produtividade, além da concentração do período de colheita em sete semanas. As coberturas plásticas do solo não influenciaram as variáveis que compõem a produtividade do tomateiro.The aim of this work was to evaluate the effect of two combinations between plant density and tip pruning height, associated to the use of plastic mulching, on plants development and fruit yield of the tomato (Duradoro hybrid grown under protected environment. Two single stem cultivation systems were compared: SC1 - tip pruning after the fifth inflorescence with 20,000 plants ha-1; SC2 - tip pruning after the second inflorescence with 50,000 plants ha-1, and three different soil cover conditions: bare soil, white polyethylene film, 50 mm-thick, and black polyethylene film, 50 mm-thick. The experimental design used was randomized block, in a split plot array. The work was carried out from June to November of 2003, at the UNESP, Ilha Solteira (São Paulo State, Brazil. The treatment SC2 increased plant height at 45 days after the transplant, and

  6. The influence of cellular seeding density in the microencapsulation of hybridoma cells.

    Science.gov (United States)

    Arús, L; Orive, G; Hernández, R; Rodriguez, A; Rojas, A; Pedraz, J L

    2005-01-01

    The aim of this study was to assess the influence of different seeding densities on the function of hybridoma cells (clone 1B5, IgG 2alpha) producing an anti-angiogenic monoclonal antibody (mAb), microencapsulated using a high-voltage electrostatic field. Viable cells were microencapsulated in alginate/poly-L-lysine/alginate (APA) capsules and maintained in tissue culture. Cellular growth rates, production and release of mAb from the capsules were assessed. This study shows that hybridoma cells survive, proliferate and remain functionally competent for over one month in vitro after microencapsulation in APA capsules generated in an electrostatic field. However, the cell seeding density had to be at least 10(7) cells/ml for the microencapsulated cells to be viable and to produce and release mAb through the capsule membrane. The maximum monoclonal antibody concentration in this culture was 29.1 microg/ml by day 17, with a tendency to increase, but capsule breakage impeded the follow-up of this determination.

  7. Significance of Mast Cell Density and Distribution in Various Histopathological Lesions of Leprosy

    Directory of Open Access Journals (Sweden)

    Santosh .L Munde

    2014-01-01

    Full Text Available Background: Leprosy also known as Hansen’s disease (HD, is a chronic infection caused by the Mycobacterium leprae. Mast cells in leprosy have been investigated in the recent past and are being examined as a basis for future studies. Materials and Methods: 119 leprosy cases and 14 control biopsies stained with toluidine blue were assessed for density and distribution of mast cells. 38 cases had undergone treatment. Results: Significantly higher mast cell count was obtained in the skin lesions of borderline lepromatous leprosy. On comparison with controls lower counts were observed in polar tuberculoid and indeterminate leprosy whereas higher values were observed in all other groups. The mast cell count in leprosy is probably determined by the pattern of cytokines released by the T lymphocytes. Mast cell density assessed after treatment was reduced in borderline tuberculoid and borderline lepromatous cases however in indeterminate group, the counts were increased. Conclusion: The distribution of mast cells in leprosy is a variable feature and there is no constant site or predilection for a particular type of leprosy.

  8. Enzymatically Modified Low-Density Lipoprotein Promotes Foam Cell Formation in Smooth Muscle Cells via Macropinocytosis and Enhances Receptor-Mediated Uptake of Oxidized Low-Density Lipoprotein.

    Science.gov (United States)

    Chellan, Bijoy; Reardon, Catherine A; Getz, Godfrey S; Hofmann Bowman, Marion A

    2016-06-01

    Enzyme-modified nonoxidized low-density lipoprotein (ELDL) is present in human atherosclerotic lesions. Our objective is to understand the mechanisms of ELDL uptake and its effects on vascular smooth muscle cells (SMC). Transformation of murine aortic SMCs into foam cells in response to ELDL was analyzed. ELDL, but not acetylated or oxidized LDL, was potent in inducing SMC foam cell formation. Inhibitors of macropinocytosis (LY294002, wortmannin, amiloride) attenuated ELDL uptake. In contrast, inhibitors of receptor-mediated endocytosis (dynasore, sucrose) and inhibitor of caveolae-/lipid raft-mediated endocytosis (filipin) had no effect on ELDL uptake in SMC, suggesting that macropinocytosis is the main mechanism of ELDL uptake by SMC. Receptor for advanced glycation end products (RAGE) is not obligatory for ELDL-induced SMC foam cell formation, but primes SMC for the uptake of oxidized LDL in a RAGE-dependent manner. ELDL increased intracellular reactive oxygen species, cytosolic calcium, and expression of lectin-like oxidized LDL receptor-1 in wild-type SMC but not in RAGE(-/-) SMC. The macropinocytotic uptake of ELDL is regulated predominantly by intracellular calcium because ELDL uptake was completely inhibited by pretreatment with the calcium channel inhibitor lacidipine in wild-type and RAGE(-/-) SMC. This is in contrast to pretreatment with PI3 kinase inhibitors which completely prevented ELDL uptake in RAGE(-/-) SMC, but only partially in wild-type SMC. ELDL is highly potent in inducing foam cells in murine SMC. ELDL endocytosis is mediated by calcium-dependent macropinocytosis. Priming SMC with ELDL enhances the uptake of oxidized LDL. © 2016 American Heart Association, Inc.

  9. Investigation of microalgae with photon density waves

    Science.gov (United States)

    Frankovitch, Christine; Reich, Oliver; Löhmannsröben, Hans-Gerd

    2007-09-01

    Phototropic microalgae have a large potential for producing valuable substances for the feed, food, cosmetics, pigment, bioremediation, and pharmacy industries as well as for biotechnological processes. Today it is estimated that the microalgal aquaculture worldwide production is 5000 tons of dry matter per year (not taking into account processed products) making it an approximately $1.25 billion U.S. per year industry. For effective observation of the photosynthetic growth processes, fast on-line sensor systems that analyze the relevant biological and technical process parameters are preferred. The optical properties of the microalgae culture influence the transport of light in the photobioreactor and can be used to extract relevant information for efficient cultivation practices. Microalgae cultivation media show a combination of light absorption and scattering, which are influenced by the concentrations and the physical and chemical properties of the different absorbing and scattering species (e.g. pigments, cell components, etc.). Investigations with frequency domain photon density waves (PDW) allow for the examination of absorption and scattering properties of turbid media, namely the absorption and reduced scattering coefficient. The reduced scattering coefficient can be used to characterize physical and morphological properties of the medium, including the cell concentration, whereas the absorption coefficient correlates with the pigment content. Nannochloropsis oculata, a single-cell species of microalgae, were examined in a nutrient solution with photon density waves. The absorption and reduced scattering coefficients were experimentally determined throughout the cultivation process, and applied to gain information about the cell concentration and average cell radius.

  10. OPTIMIZATION OF Chlorella vulgaris BEIJ. CULTIVATION IN A BIOREACTOR OF CONTINUOUS ACTION

    Directory of Open Access Journals (Sweden)

    Bodnar

    2016-08-01

    Full Text Available The aim of the research was the development and testing of bioreactor for intensive cultivation of algae Chlorella vulgaris Beij. with fully controlled conditions within the operating parameters according to the selected evaluation criteria of the cultivation process. To check the functioning efficiency of the decigned photobioreactor, the growth of Chlorella vulgaris Beij. (CHLOROPHYTA in Fitzgerald’s medium modified by Zender and Gorham (No 11 under the artificial illumination with daylight electric lamps (intensity of 2 500 Lx for 16 hours a day at 22–25 ºС was studed. It was found that at stabilization of culture conditions the maximum value of culture density was observed at the 18th day of cultivation. In this moment, the amount of cells reached 269.2 ± 3.0·109 cells/l, while cells amount in stationary phase was within 110.1 ± 4.9·109 cells/l. This makes it possible the continuous chlorella cultivation with an average productivity in stationary mode of about 110 ± 4 mg/l of dry mass with protein content about 60 mg, carbohydrates about 35 and lipids about 12 mg of dry mass/l. Sunlight and activators of biosynthesis of organic substances of individual classes allow changeing the ratio of proteins, carbohydrates and lipids that is prospective for further research.

  11. Results of Current Density Distribution Mapping in PEM Fuel Cells Dependent on Operation Parameters

    Directory of Open Access Journals (Sweden)

    Zbigniew A. Styczynski

    2013-07-01

    Full Text Available This paper presents in situ measurements of a newly developed current density measurement system for proton exchange membrane fuel cells (PEMFC. While the functional principle and technical evaluation of the measurement system were presented in a previous paper, this paper analyzes the influence of various operation parameters, including multiple start-stop operation, at the anode, cathode and cooling locations on the distribution and long-term development of the current density. The system was operated for 500 h over two years with long periods of inactivity between measurements. The measurement results are evaluated and provide additional information on how to optimize the operation modes of fuel cells, including the start and stop of such systems as well as the water balance.

  12. High cell density strategy for poly(3-hydroxybutyrate production by Cupriavidus necator

    Directory of Open Access Journals (Sweden)

    J. L. Ienczak

    2011-12-01

    Full Text Available Poly(3-hydroxybutyrate (P(3HB is a carbon and intracellular storage source for different microorganisms and its production can achieve high productivities by means of high cell density cultures. The aim of this study was to propose a high cell density strategy for P(3HB production by Cupriavidus necator. The exponential growth phase demands an accurate control of the oxygen transfer system in the bioreactor, due to maximum specific growth rate (µXr, and, consequently, a maximum specific oxygen uptake rate (QO2, in addition to significant residual biomass (Xr growth in high cell density cultures. In this context, this work investigated the strategy for obtaining high cell density, with the inclusion of a linear growth phase for P(3HB production by C. necator in a fed-batch culture. The linear growth phase was included between the exponential growth phase and the P(3HB production phase as a strategy to reduce the specific growth rate (µXr and specific oxygen uptake rate (QO2, with constant residual biomass growth rate (d(V.Xr/dt = k = constant and linear increase of biomass. Three strategies of culture were performed. The results showed that a high residual biomass concentration (30 gXr.L-1 can be reached by the inclusion of the linear growth strategy and specific growth rates (µXr between 0.08 and 0.05 h-1, at the beginning of the production phase, are necessary to attain a high P(3HB productivity.

  13. Best practices in heterotrophic high-cell-density microalgal processes: achievements, potential and possible limitations

    OpenAIRE

    Bumbak, Fabian; Cook, Stella; Zachleder, Vilém; Hauser, Silas; Kovar, Karin

    2011-01-01

    Microalgae of numerous heterotrophic genera (obligate or facultative) exhibit considerable metabolic versatility and flexibility but are currently underexploited in the biotechnological manufacturing of known plant-derived compounds, novel high-value biomolecules or enriched biomass. Highly efficient production of microalgal biomass without the need for light is now feasible in inexpensive, well-defined mineral medium, typically supplemented with glucose. Cell densities of more than 100 g l−1...

  14. "Bleaching" glycerol in a microfluidic fuel cell to produce high power density at minimal cost.

    Science.gov (United States)

    Martins, Cauê A; Ibrahim, Omar A; Pei, Pei; Kjeang, Erik

    2018-01-07

    Glycerol/bleach flow-through microfluidic fuel cells are presented. Carbon paper-modified Pt/C nanoparticles were used as the anode and cathode. Glycerol oxidation in alkaline medium was tested against hypochlorite reduction in alkaline and acidic media. The mixed media system displayed a power density of 315 mW cm-2 and an open circuit voltage of 1.9-2.0 V.

  15. Systems biology. Conditional density-based analysis of T cell signaling in single-cell data.

    Science.gov (United States)

    Krishnaswamy, Smita; Spitzer, Matthew H; Mingueneau, Michael; Bendall, Sean C; Litvin, Oren; Stone, Erica; Pe'er, Dana; Nolan, Garry P

    2014-11-28

    Cellular circuits sense the environment, process signals, and compute decisions using networks of interacting proteins. To model such a system, the abundance of each activated protein species can be described as a stochastic function of the abundance of other proteins. High-dimensional single-cell technologies, such as mass cytometry, offer an opportunity to characterize signaling circuit-wide. However, the challenge of developing and applying computational approaches to interpret such complex data remains. Here, we developed computational methods, based on established statistical concepts, to characterize signaling network relationships by quantifying the strengths of network edges and deriving signaling response functions. In comparing signaling between naïve and antigen-exposed CD4(+) T lymphocytes, we find that although these two cell subtypes had similarly wired networks, naïve cells transmitted more information along a key signaling cascade than did antigen-exposed cells. We validated our characterization on mice lacking the extracellular-regulated mitogen-activated protein kinase (MAPK) ERK2, which showed stronger influence of pERK on pS6 (phosphorylated-ribosomal protein S6), in naïve cells as compared with antigen-exposed cells, as predicted. We demonstrate that by using cell-to-cell variation inherent in single-cell data, we can derive response functions underlying molecular circuits and drive the understanding of how cells process signals. Copyright © 2014, American Association for the Advancement of Science.

  16. Evaluation of Mast Cell and Blood Vessel Density in Inflammatory Periapical Lesions

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    Safoura Seifi

    2012-01-01

    Full Text Available Introduction: Radicular cystsand periapical granulomas are the most common periapical inflammatory lesions. However, the role of cellular immunity and microvessels in their pathogenesis remains unknown. The aim of this study was to evaluate the mast cell density (MCD, mircovessel density (MVD and investigating the correlation between their densities with each other in the above mentioned lesions.Materials & Methods: In this descriptive cross-sectional study, 40 paraffin blocks of mentioned lesions were selected from achieves of School of Dentistry, Babol University of Medical Sciences. Three sections were prepared from each block and stained by hematoxylin-eosin, toluidine blue, and immunohistochemically for CD34 to determine the score of inflammation, presence of mast cells and degranulatedmast cells (DMCs, and MVD, respectively. The correlation between MCD and either inflammatory infiltrate or MVD was evaluated. Data analyzed by t student, Mann-Whitney and Spearman test.Results: Mast cells were present in all periapical inflammatory lesions; 15.4±14.8 for MCD, 7.2±6.1 for DMCs, and the ratio of DMCs to total number of MCs was 0.354±0.166 and 14.8+4.44 for blood vessel density in radicular cyst and 8.52±6.75, 2.91±2.1, 0.196±0.194 and 13±8.02 in periapical granulomas, respectively. There was a positive correlation between MCD and MVD in radicular cyst (P=0.03, r=0.341, but not in periapical granulomas (P=0.6, r=0.124. MCD and MVD increased with the score of inflammation in radicular cyst (P=0.001, r=0.7 and periapical granuloma (P=0.012, r=0.54.Conclusion: Mast cells and microvessels play a role in pathogenesis of periapical inflammatory lesions. In this study, the density of mast cells and DMCs in radicular cyst was higher than periapical granulomas, but no difference was observed regarding MVD in periapical inflammatory lesions. It seems that the relationship between MCD and MVD is different based on the clinical stage of periapical

  17. Microvascular density, CD68 and tryptase expression in human diffuse large B-cell lymphoma.

    Science.gov (United States)

    Marinaccio, Christian; Ingravallo, Giuseppe; Gaudio, Francesco; Perrone, Tommasina; Nico, Beatrice; Maoirano, Eugenio; Specchia, Giorgina; Ribatti, Domenico

    2014-11-01

    Diffuse Large B-cell Lymphoma (DLBCL) is the most common form of Non-Hodgkin lymphoma characterized by clinical and biological heterogeneity attributable both to the tumor cells and the complex tumor-microenvironment surrounding them. Tumor-associated macrophages (TAMs) and mast cells are two major components of the tumor inflammatory infiltrate with a definite role in enhancing tumor angiogenesis. In this study, we have investigated CD68 and tryptase expression and their relationship with microvascular density (MVD) in chemo-resistant and chemosensitive patients affected by DLBCL. CD68 and tryptase expression as well as MVD were increased in chemo-resistant patients when compared with chemosensitive patients. Tryptase expression showed a positive correlation with MVD, supporting a role for mast cell in DLBCL tumor angiogenesis, while CD68 correlation with MVD was not significant, indicating a different role for TAMs than angiogenesis in DLBCL. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Low density lipoprotein inhibits accumulation of nitrites in murine brain endothelial cell cultures.

    Science.gov (United States)

    Bereta, M; Bereta, J; Cohen, S; Cohen, M C

    1992-07-15

    Endothelial cells produce nitric oxide which is considered to serve as a major source of endothelial derived relaxing factor activity. It has been demonstrated that activation of mouse brain endothelium by TNF-alpha and IFN-gamma led to accumulation of nitrite which is presumably formed by oxidation of nitric oxide. A number of studies suggest that reactive oxygen species produced by cytokine-activated cells are involved in the conversion of nitric oxide to nitrites and nitrates. We investigated whether low density lipoprotein (LDL), acting as a radical scavenger, is able to inhibit nitrite accumulation in mouse brain endothelial cell cultures and in a cell-free system in which sodium nitroprusside was used as a source of nitric oxide. A comparison of these two models indicates the active involvement of LDL in suppressing nitrite accumulation in murine endothelial cultures.

  19. Relationship of mast cell density with lymphangiogenesis and prognostic parameters in breast carcinoma.

    Science.gov (United States)

    Keser, Sevinc H; Kandemir, Nilufer O; Ece, Dilek; Gecmen, Gonca G; Gul, Aylin E; Barisik, Nagehan O; Sensu, Sibel; Buyukuysal, Cagatay; Barut, Figen

    2017-04-01

    In many cancers, mast cell density (MCD) in the tumor microenvironment is associated with tumor progression and, to a greater extent, angiogenesis. Our study was designed to investigate the correlation between MCD, tumor lymphangiogenesis, and several well-established prognostic parameters in breast cancer. One hundred and four cases of invasive breast carcinoma diagnosed in our clinic between 2007 and 2011 were included. Mast cells and lymphatic vessels were stained with toluidine blue and D2-40, respectively, and their densities were calculated in various areas of tumors and lymph nodes. The variables of MCD and lymphatic vessel density (LVD) were compared using prognostic parameters as well as with each other. As tumor size and volume increased, MCD increased comparably in metastatic lymph nodes; intratumoral and peritumoral LVD also increased. Lymphovascular invasion, lymphatic invasion, perineural invasion, and estrogen receptor positivity were positively related to intratumoral MCD. The relationship between peritumoral MCD and nontumoral breast tissue MCD was statistically significant. Stage was correlated with MCD in metastatic lymph nodes. Metastatic lymph node MCD and intratumoral MCD were also significantly related. Stage, lymphatic invasion, perineural invasion, lymphovascular invasion, and metastatic lymph node MCD were all correlated with intratumoral and/or peritumoral LVD. As nuclear grade increased, intratumoral LVD became higher. In breast carcinoma, MCD, depending on its location, was related to several prognostic parameters. Notably, mast cells may have at least some effect on lymphangiogenesis, which appears to be a predictor of tumor progression. Copyright © 2017. Published by Elsevier Taiwan.

  20. A Semianalytical Model Using MODIS Data to Estimate Cell Density of Red Tide Algae (Aureococcus anophagefferens

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    Lingling Jiang

    2016-01-01

    Full Text Available A multiband and a single-band semianalytical model were developed to predict algae cell density distribution. The models were based on cell density (N dependent parameterizations of the spectral backscattering coefficients, bb(λ, obtained from in situ measurements. There was a strong relationship between bb(λ and N, with a minimum regression coefficient of 0.97 at 488 nm and a maximum value of 0.98 at other bands. The cell density calculated by the multiband inversion model was similar to the field measurements of the coastal waters (the average relative error was only 8.9%, but it could not accurately discern the red tide from mixed pixels, and this led to overestimation of the area affected by the red tide. While the single-band inversion model is less precise than the former model in the high chlorophyll water, it could eliminate the impact of the suspended sediments and make more accurate estimates of the red tide area. We concluded that the two models both have advantages and disadvantages; these methods lay the foundation for developing a remote sensing forecasting system for red tides.

  1. Relationship of mast cell density with lymphangiogenesis and prognostic parameters in breast carcinoma

    Directory of Open Access Journals (Sweden)

    Sevinc H. Keser

    2017-04-01

    Full Text Available In many cancers, mast cell density (MCD in the tumor microenvironment is associated with tumor progression and, to a greater extent, angiogenesis. Our study was designed to investigate the correlation between MCD, tumor lymphangiogenesis, and several well-established prognostic parameters in breast cancer. One hundred and four cases of invasive breast carcinoma diagnosed in our clinic between 2007 and 2011 were included. Mast cells and lymphatic vessels were stained with toluidine blue and D2-40, respectively, and their densities were calculated in various areas of tumors and lymph nodes. The variables of MCD and lymphatic vessel density (LVD were compared using prognostic parameters as well as with each other. As tumor size and volume increased, MCD increased comparably in metastatic lymph nodes; intratumoral and peritumoral LVD also increased. Lymphovascular invasion, lymphatic invasion, perineural invasion, and estrogen receptor positivity were positively related to intratumoral MCD. The relationship between peritumoral MCD and nontumoral breast tissue MCD was statistically significant. Stage was correlated with MCD in metastatic lymph nodes. Metastatic lymph node MCD and intratumoral MCD were also significantly related. Stage, lymphatic invasion, perineural invasion, lymphovascular invasion, and metastatic lymph node MCD were all correlated with intratumoral and/or peritumoral LVD. As nuclear grade increased, intratumoral LVD became higher. In breast carcinoma, MCD, depending on its location, was related to several prognostic parameters. Notably, mast cells may have at least some effect on lymphangiogenesis, which appears to be a predictor of tumor progression.

  2. Effects of dissolved oxygen level on cell growth and total lipid accumulation in the cultivation of Rhodotorula glutinis.

    Science.gov (United States)

    Yen, Hong-Wei; Zhang, Zhiyong

    2011-07-01

    The total amount of lipids produced in Rhodotorula glutinis is a subject which has attracted increasing attention due to the potential biodiesel conversion from these microbial oils. The effects of the dissolved oxygen (DO) level in lipid accumulation were examined in this study. Variations of different medium volumes (30, 40 and 50ml) and shaking speed (60, 150 and 210rpm) in the flask trials were adopted to explore the DO effects on lipid production. All of the results revealed that a low DO could retard cell growth, while enhancing lipid accumulation. The 5l-fermentor results also confirm that a low DO (25 ± 10%) batch could have higher lipid content than that of high DO batch (60 ± 10%). Nevertheless, the DO level would not obviously affect the lipid composition profile. Oleic acid (C18:1) was the primary fatty acid in both batches. Due to the slow biomass growth rate resulting from the low DO, a two-stage DO controlled strategy (consisting of a high DO stage and following a low DO stage) was performed to improve the cell growth and lipid accumulation simultaneously. However, the strategy was not successful on the enhancement of total lipid production as compared to other batches. Conclusively, even a low DO could retard cell growth; the total production of lipids in the batch with low DO was higher that of the high DO batch due to the enhancement of lipid accumulation. Therefore, the batch operation of R. glutinis at the low DO was suggested for the purpose of lipid production. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  3. Comparison of clinical grade human platelet lysates for cultivation of mesenchymal stromal cells from bone marrow and adipose tissue

    DEFF Research Database (Denmark)

    Juhl, Morten; Tratwal, Josefine; Follin, Bjarke

    2016-01-01

    different commercially available hPL fulfilling good manufacturing practice criteria for clinical use. BMSCs and ASCs cultured in Minimum Essential Medium Eagle-alpha supplemented with 5% PLT-Max (Mill Creek), Stemulate™ PL-S and Stemulate™ PL-SP (COOK General Biotechnology) were compared to standard...... culture conditions with 10% fetal bovine serum (FBS). Cell morphology, proliferation, phenotype, genomic stability, and differentiation potential were analyzed. RESULTS: Regardless of manufacturer, BMSCs and ASCs cultured in hPL media showed a significant increase in proliferation capacity compared to FBS...

  4. The role of cell size in density gradient electrophoretic separation of mouse leukemia cells according to position in the cell cycle

    Science.gov (United States)

    Plank, L. D.; Kunze, M. E.; Todd, P. W.

    1985-01-01

    Cultured mouse leukemia cells line L5178Y were subjected to upward electrophoresis in a density gradient and the slower migrating cell populations were enriched in G2 cells. It is indicated that this cell line does not change electrophoretic mobility through the cell cycle. The possibility that increased sedimentation downward on the part of the larger G2 cells caused this separation was explored. Two different cell populations were investigated. The log phase population was found to migrate upward faster than the G2 population, and a similar difference between their velocities and calculated on the basis of a 1 um diameter difference between the two cell populations. The G2 and G1 enriched populations were isolated by Ficoll density gradient sedimentation. The bottom fraction was enriched in G2 cells and the top fraction was enriched with G1 cells, especially when compared with starting materials. The electrophoretic mobilities of these two cell populations did not differ significantly from one another. Cell diameter dependent migration curves were calculated and were found to be different. Families of migration curves that differ when cell size is considered as a parameter are predicted.

  5. Effect of purified fractions from cell culture supernate of high-density pre-B acute lymphoblastic leukemia cells (ALL3) on the growth of ALL3 cells at low density.

    Science.gov (United States)

    Patel, Sapan J; Darie, Costel C; Clarkson, Bayard D

    2017-02-01

    The mechanisms underlying the aberrant growth and interactions between cells are not understood very well. The pre-B acute lymphoblastic leukemia cells directly obtained from an adult patient grow very poorly or do not grow at all at low density (LD), but grow better at high starting cell density (HD). We found that the LD ALL3 cells can be stimulated to grow in the presence of diffusible, soluble factors secreted by ALL3 cells themselves growing at high starting cell density. We then developed a biochemical purification procedure that allowed us to purify the factor(s) with stimulatory activity and analyzed them by nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS). Using nanoLC-MS/MS we have identified several proteins which were further processed using various bioinformatics tools. This resulted in eight protein candidates which might be responsible for the growth activity on non-growing LD ALL3 cells and their involvement in the stimulatory activity are discussed. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Current Density Distribution Mapping in PEM Fuel Cells as An Instrument for Operational Measurements

    Directory of Open Access Journals (Sweden)

    Martin Geske

    2010-04-01

    Full Text Available A newly developed measurement system for current density distribution mapping has enabled a new approach for operational measurements in proton exchange membrane fuel cells (PEMFC. Taking into account previously constructed measurement systems, a method based on a multi layer printed circuit board was chosen for the development of the new system. This type of system consists of a sensor, a special electronic device and the control and visualization PC. For the acquisition of the current density distribution values, a sensor device was designed and installed within a multilayer printed circuit board with integrated shunt resistors. Varying shunt values can be taken into consideration with a newly developed and evaluated calibration method. The sensor device was integrated in a PEM fuel cell stack to prove the functionality of the whole measurement system. A software application was implemented to visualize and save the measurement values. Its functionality was verified by operational measurements within a PEMFC system. Measurement accuracy and possible negative reactions of the sensor device during PEMFC operation are discussed in detail in this paper. The developed system enables operational measurements for different operating phases of PEM fuel cells. Additionally, this can be seen as a basis for new opportunities of optimization for fuel cell design and operation modes.

  7. Controlling Side Chain Density of Electron Donating Polymers for Improving VOC in Polymer Solar Cells

    Science.gov (United States)

    Kim, B. J.; Kim, K. H.; Cho, C. H.; Kang, H.; Yoon, S. C.

    2012-02-01

    The ability to tune the LUMO/HOMO levels of electroactive materials in active layer of polymer solar cells is critical in controlling their optical and electrochemical properties because the HOMO and LUMO offsets between the polymer donor and the electron acceptor strongly affect charge separation and the open circuit voltage (VOC) of a solar cell. Here, we developed two series of electroactive materials for improving VOC in polymer solar cells. First, we enable facile control over the number of solubilizing groups ultimately tethered to the fullerene by tuning the molar ratio between reactants from 1:1 to 1:3, thus producing o-xylenyl C60 mono-, bis-, and tris-adducts (OXCMA, OXCBA, and OXCTA) as electron acceptors with different LUMO levels. As the number of solubilizing groups increased, VOC values of the P3HT-based BHJ solar cells increased from 0.63, 0.83, to 0.98 V. Second, we present a series of novel poly[3-(4-n-octyl)phenylthiophene] (POPT) derivatives (POPT, POPTT, and POTQT) as electron donors with different side-chain density. As a result of lower HOMO levels by decrease in the side-chain density of the polymers, the devices consisting of POPT, POPTT, and POPQT with PCBM showed increased VOC values of 0.58, 0.63, and 0.75 V, respectively.

  8. Effects of Temperature variations on the Super Fine Powderization of Korean Cultivated Wild Ginseng

    Directory of Open Access Journals (Sweden)

    Jin Ho Kim

    2006-12-01

    Full Text Available Objectives : The aim of this study was to find optimal conditions for producing red ginseng from cultivated wild ginseng using the Turbo Mill. Methods : Characteristics of powdered cultivated wild ginseng based on various temperature settings of the Turbo Mill were observed, and changes in the content was measured by HPLC for various ginsenosides. Results : 1. The diameter of cultivated wild ginseng powder ground by the Turbo Mill was around 10㎛. 2. As the temperature rose, presusre, Specific Mechanical Energy(SME, and density decreased, whileas Water Solubility Index(WSI increased. 3. As the temperature rose, super fine powder showed tendency to turn into dark brown. 4. Measuring content changes by HPLC, there was no detection of ginsenoside Rg3 and ginsenosideRg1, Rb1, and Rh2 concentrations decreased with increase in temperature. Conclusions : Super fine powder of cultivated wild ginseng produced by the Turbo Mill promotes easy absorption of effective ingredients by breaking the cell walls. Using this mechanism to produce red ginseng from cultivated wild ginseng, it yielded less than satisfactory results under the current experiment setup. Furtherresearches are needed to verify more suitable condition for the production of red ginseng.

  9. Comparative analysis of two colorimetric assays in dental pulp cell density

    Science.gov (United States)

    Nedel, F.; Soki, F.N.; Conde, M.C.M.; Zeitlin, B.D.; Tarquinio, S.B.C.; Nör, J.E.; Seixas, F.K.; Demarco, F.F.

    2013-01-01

    Aim To compare and contrast two colorimetric assays used for measurement of proliferation using two dental pulp cell types: dental pulp stem cells (DPSC) and human dental pulp fibroblasts (HDPF). Methodology DPSC or HDPF were seeded at 0.25 × 104 cells/well in 96-well plates. Cell proliferation was evaluated after 24-72 hours. At the end of the experimental period, the Sulforhodamine B (SRB) assay or a water-soluble tetrazolium salt (WST-1) assay was performed. Optical densities were determined in a microplate reader (Genius; TECAN). Data were analyzed by Student’s t-test (comparison between cell types), and one-way ANOVA followed by Tukey test (time point intervals). Pearson’ correlation tests were performed to compare the two assays for each cell line. Results Both assays showed that DPSC had higher proliferation rates than HDPF. A positive significant correlation between the two colorimetric assays tested for both cell types DPSC (Pearson’s Correlation Coefficient=0.847; passay is therefore one of practical application. SRB stained plates may be dried and stored so may have utility in laboratories where data may require review or when access to analytical equipment is limited. WST-1 assays have the benefit of both ease and speed and may have utility in laboratories requiring either high throughput or rapid analyses. PMID:20880134

  10. Densidade de um planossolo sob sistemas de cultivo avaliada por meio da tomografia computadorizada de raios gama Bulk density of an alfisol under cultivation systems in a long-term experiment evaluated with gamma ray computed tomography

    Directory of Open Access Journals (Sweden)

    Adilson Luís Bamberg

    2009-10-01

    lowland soils is based on the use of crop rotation and succession, which are essential for the control of red and black rice. The effects on the soil properties deserve studies, particularly on soil compaction. The objective of this study was to identify compacted layers in an Albaqualf under different cultivation and tillage systems, by evaluating the soil bulk density (Ds with Gamma Ray Computed Tomography (TC. The analysis was carried out in a long-term experiment, from 1985 to 2004, at an experimental station of Embrapa Clima Temperado, Capão do Leão, RS, Brazil, in a random block design with seven treatments, with four replications (T1 - one year rice with conventional tillage followed by two years fallow; T2 - continuous rice under conventional tillage; T4 - rice and soybean (Glycine Max L. rotation under conventional tillage; T5 - rice, soybean and corn (Zea maize L. rotation under conventional tillage; T6 - rice under no-tillage in the summer in succession to rye-grass (Lolium multiflorum L. in the winter; T7 - rice under no-tillage and soybean under conventional tillage rotation; T8 - control: uncultivated soil. The Gamma Ray Computed Tomography method did not identify compacted soil layers under no-tillage rice in succession to rye-grass; two fallow years in the irrigated rice production system did not prevent the formation of a compacted layer at the soil surface; and in the rice, soybean and corn rotation under conventional tillage two compacted layers were identified (0.0 to 1.5 cm and 11 to 14 cm, indicating that they may restrict the agricultural production in this cultivation system on Albaqualf soils.

  11. Enhanced photocurrent density of HTM-free perovskite solar cells by carbon quantum dots

    Science.gov (United States)

    Zou, Haiyuan; Guo, Daipeng; He, Bowen; Yu, Jiaguo; Fan, Ke

    2018-02-01

    Full-printable and hole transport material (HTM)-free perovskite solar cells (PSCs) with carbon counter electrodes feature high stability and low cost. However, the perovskite film prepared by conventional one-step solution-coating method always shows a relatively poor coverage on the substrate, leading to the limit of the photocurrent density. In this study, we incorporated carbon quantum dots (CQDs) in the perovskite films, and investigated their effects on the performance of TiO2 nanosheet-based and HTM-free PSCs. It was found that the addition of CQDs to the perovskite film can enhance the photocurrent density of PSCs, and the optimal PSC with 0.1% CQDs evolved 60% higher photocurrent density than the pristine one. The improved photocurrent density was attributed to the heterogeneous nuclei derived from CQDs during perovskite crystallization, which would increase amount of perovskite nuclei and form a fine perovskite grain, leading to a better coverage on the substrate. Moreover, due to the excellent conductivity, CQDs in perovskite films could efficiently transport the photo-excited electrons, accelerating the separation and mobilization of charge carriers. This study presents the incorporation of CQDs in perovskite as an efficient approach to promote the performance of HTM-free PSCs.

  12. Detection of Toxoplasma gondii in raw caprine, ovine, buffalo, bovine, and camel milk using cell cultivation, cat bioassay, capture ELISA, and PCR methods in Iran.

    Science.gov (United States)

    Dehkordi, Farhad Safarpoor; Borujeni, Mohammad Reza Haghighi; Rahimi, Ebrahim; Abdizadeh, Rahman

    2013-02-01

    This study was conducted to determine the presence of Toxoplasma gondii in animal milk samples in Iran. From a total of 395 dairy herds in three provinces of Iran, 66 bovine, 58 ovine, 54 caprine, 33 buffalo, and 30 camel herds were studied, and from these parts of Iran, 200 bovine, 185 ovine, 180 caprine, 164 buffalo, and 160 camel milk samples were collected from various seasons. Samples were tested for Toxoplasma gondii by cell line culture, enzyme-linked immunosorbent assay (ELISA), and polymerase chain reaction (PCR) technique. Only the results of cell line cultivation were confirmed by bioassay in cat. Results indicated that all herds were infected with Toxoplasma gondii. The culture method showed that 51 out of 889 milk samples (5.73%) were positive for Toxoplasma gondii, and all 51 positive culture results were positive with bioassay in cat. The Fars province had the highest prevalence of Toxoplasma gondii (6.84%). The ELISA test showed that 41 milk samples (4.61%) were positive for the presence of Toxoplasma gondii, while the PCR showed that 46 milk samples were positive for Toxoplasma gondii. The results showed higher sensitivity of PCR and higher specificity of ELISA. Caprine had the highest (10%) and camel had the lowest (3.12%) prevalence rate of parasite. The summer season had the highest (76.47%) but winter (3.92) had the lowest incidence of Toxoplasma gondii. This study is the first prevalence report of direct detection of Toxoplasma gondii in animal milk samples in Iran.

  13. Assessment of corneal endothelial cell density in patients with keratoconus not using contact lenses.

    Science.gov (United States)

    Timucin, Ozgur Bulent; Karadag, Mehmet Fatih; Cinal, Adnan; Asker, Muntecep; Asker, Selvi; Timucin, Damla

    2013-04-01

    To assess the corneal endothelial cell density (ECD) in keratoconus patients with no history of contact lens use. Yuzuncu Yil University, School of Medicine and Van Training and Research Hospital, Department of Ophthalmology, Van, Turkey. Cross-sectional controlled study. The eyes of 65 patients with the diagnosis of keratoconus with no history of contact lens wear and the eyes of 40 healthy controls were prospectively examined using the Heidelberg Retinal Tomography Rostock Cornea Module (HRT3/RCM). The average ECD from the two groups were then compared. Of the cases with keratoconus, 44 (67.7%) were men and 21 (32.3%) were women. The mean age was 20.9±6.8 (range=10-41) years. Of the controls, 28 (70%) were men and 12 (30%) were women. The mean age was 23.9±5.8 (range=14-35) years. Of the 65 eyes with keratoconus, 19 (29.2%) had mild keratoconus, 21 (32.3%) had moderate keratoconus, and 25 (38.5%) had severe keratoconus. The mean ECD was 2731.6±303.2 cells/mm2 in cases with keratoconus and 2664.9±319.5 cells/mm2 in controls. There was no difference between the densities (unpaired t-test, P=0.4). No significant relationships were found between the ECD data and central corneal thickness or steepest keratometric. Endothelial cell density was unaltered in keratoconic patients without a history of contact lens use when compared with healthy controls. Change in ECD is independent from the central corneal thickness and the stage of keratoconus. Copyright © 2012 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

  14. Plasma membrane temperature gradients and multiple cell permeabilization induced by low peak power density femtosecond lasers

    Directory of Open Access Journals (Sweden)

    Allen L. Garner

    2016-03-01

    Full Text Available Calculations indicate that selectively heating the extracellular media induces membrane temperature gradients that combine with electric fields and a temperature-induced reduction in the electropermeabilization threshold to potentially facilitate exogenous molecular delivery. Experiments by a wide-field, pulsed femtosecond laser with peak power density far below typical single cell optical delivery systems confirmed this hypothesis. Operating this laser in continuous wave mode at the same average power permeabilized many fewer cells, suggesting that bulk heating alone is insufficient and temperature gradients are crucial for permeabilization. This work suggests promising opportunities for a high throughput, low cost, contactless method for laser mediated exogenous molecule delivery without the complex optics of typical single cell optoinjection, for potential integration into microscope imaging and microfluidic systems.

  15. Low-density open-cell foams in the NiTi system

    Science.gov (United States)

    Grummon, David S.; Shaw, John A.; Gremillet, Antoine

    2003-04-01

    It is shown that open-cell metallic foams having very low density, and that display martensite transformations required for shape memory and superelastic behavior, can be fabricated using a powder-metallurgy technique. Results are presented on experiments in which a polymeric precursor foam was coated with an equiatomic NiTi powder slurry and subsequently sintered to yield foams with relative densities as low as 0.039. Although contaminated with interstitial impurities, they displayed unambiguous calorimetric signature of the B2→B19' transformation. The results are of considerable significance to potential applications requiring ultralightweight structures with the unusual dissipative and strain-recovery properties of NiTi shape-memory materials.

  16. Analysis of electronic models for solar cells including energy resolved defect densities

    Energy Technology Data Exchange (ETDEWEB)

    Glitzky, Annegret

    2010-07-01

    We introduce an electronic model for solar cells including energy resolved defect densities. The resulting drift-diffusion model corresponds to a generalized van Roosbroeck system with additional source terms coupled with ODEs containing space and energy as parameters for all defect densities. The system has to be considered in heterostructures and with mixed boundary conditions from device simulation. We give a weak formulation of the problem. If the boundary data and the sources are compatible with thermodynamic equilibrium the free energy along solutions decays monotonously. In other cases it may be increasing, but we estimate its growth. We establish boundedness and uniqueness results and prove the existence of a weak solution. This is done by considering a regularized problem, showing its solvability and the boundedness of its solutions independent of the regularization level. (orig.)

  17. Modeling quorum sensing trade-offs between bacterial cell density and system extension from open boundaries

    Science.gov (United States)

    Marenda, Mattia; Zanardo, Marina; Trovato, Antonio; Seno, Flavio; Squartini, Andrea

    2016-12-01

    Bacterial communities undergo collective behavioural switches upon producing and sensing diffusible signal molecules; a mechanism referred to as Quorum Sensing (QS). Exemplarily, biofilm organic matrices are built concertedly by bacteria in several environments. QS scope in bacterial ecology has been debated for over 20 years. Different perspectives counterpose the role of density reporter for populations to that of local environment diffusivity probe for individual cells. Here we devise a model system where tubes of different heights contain matrix-embedded producers and sensors. These tubes allow non-limiting signal diffusion from one open end, thereby showing that population spatial extension away from an open boundary can be a main critical factor in QS. Experimental data, successfully recapitulated by a comprehensive mathematical model, demonstrate how tube height can overtake the role of producer density in triggering sensor activation. The biotic degradation of the signal is found to play a major role and to be species-specific and entirely feedback-independent.

  18. Distribution of the Current Density in Electrolyte of the Pem Fuel Cell

    Directory of Open Access Journals (Sweden)

    Eugeniusz Kurgan

    2004-01-01

    Full Text Available In this paper water management in proton exchange membrane (PEM fuel cell is considered. Firt mass convervation law for water is applied. Next proton transport is described by the Nernst-Planck equation and liqid water convection velocity is eliminated by the Schlogl equation. Electro-osmotic drag coefficient is related to hydrogen index and experimentally determined swelling coefficient. Three partial differential equations for molar water concentration Cw, electric potential ϕ and water pressure Pw are formulated. Current density vector i is derived from proton flux expression. These equations together with adequate boundary conditions were solved using finite element method. The distribution of electric potential and current density in function of geometrical parametres is investigated. At the end some illustrative example is given.

  19. Modeling quorum sensing trade-offs between bacterial cell density and system extension from open boundaries.

    Science.gov (United States)

    Marenda, Mattia; Zanardo, Marina; Trovato, Antonio; Seno, Flavio; Squartini, Andrea

    2016-12-14

    Bacterial communities undergo collective behavioural switches upon producing and sensing diffusible signal molecules; a mechanism referred to as Quorum Sensing (QS). Exemplarily, biofilm organic matrices are built concertedly by bacteria in several environments. QS scope in bacterial ecology has been debated for over 20 years. Different perspectives counterpose the role of density reporter for populations to that of local environment diffusivity probe for individual cells. Here we devise a model system where tubes of different heights contain matrix-embedded producers and sensors. These tubes allow non-limiting signal diffusion from one open end, thereby showing that population spatial extension away from an open boundary can be a main critical factor in QS. Experimental data, successfully recapitulated by a comprehensive mathematical model, demonstrate how tube height can overtake the role of producer density in triggering sensor activation. The biotic degradation of the signal is found to play a major role and to be species-specific and entirely feedback-independent.

  20. A Methodological Investigation of Cultivation.

    Science.gov (United States)

    Rubin, Alan M.; And Others

    Cultivation theory states that television engenders negative emotions in heavy viewers. Noting that cultivation methodology contains an apparent response bias, a study examined relationships between television exposure and positive restatements of cultivation concepts and tested a more instrumental media uses and effects model. Cultivation was…

  1. Plasmin promotes foam cell formation by increasing macrophage catabolism of aggregated low-density lipoprotein.

    Science.gov (United States)

    Haka, Abigail S; Grosheva, Inna; Singh, Rajesh K; Maxfield, Frederick R

    2013-08-01

    The plasmin/plasminogen system is involved in atherosclerosis. However, the mechanisms by which it stimulates disease are not fully defined. A key event in atherogenesis is the deposition of low-density lipoprotein (LDL) on arterial walls where it is modified, aggregated, and retained. Macrophages are recruited to clear the lipoproteins, and they become foam cells. The goal of this study was to assess the role of plasmin in macrophage uptake of aggregated LDL and foam cell formation. Plasminogen treatment of macrophages catabolizing aggregated LDL significantly accelerated foam cell formation. Macrophage interaction with aggregated LDL increased the surface expression of urokinase-type plasminogen activator receptor and plasminogen activator activity, resulting in increased ability to generate plasmin at the cell surface. The high local level of plasmin cleaves cell-associated aggregated LDL, allowing a portion of the aggregate to become sequestered in a nearly sealed, yet extracellular, acidic compartment. The low pH in the plasmin-induced compartment allows lysosomal enzymes, delivered via lysosome exocytosis, greater activity, resulting in more efficient cholesteryl ester hydrolysis and delivery of a large cholesterol load to the macrophage, thereby promoting foam cell formation. These findings highlight a critical role for plasmin in the catabolism of aggregated LDL by macrophages and provide a new context for considering the atherogenic role of plasmin.

  2. Non-invasive optoacoustic probing of the density and stiffness of single biological cells

    Science.gov (United States)

    Dehoux, T.; Audoin, B.

    2012-12-01

    Recently, the coherent generation of GHz acoustic waves using ultrashort laser pulses has demonstrated the ability to probe the sound velocity in vegetal cells and in cell-mimicking soft micro-objects with micrometer resolution, opening tremendous potentialities for single-cell biology. However, manipulating biological media in physiological conditions is often a technical challenge when using a laser-based setup. In this article, we present a new opto-acoustic bio-transducer composed of a thin metal film sputtered on a transparent heat sink that allows reducing importantly the laser-induced cellular stresses, and offers a wide variety of optical configurations. In particular, by exploiting the acoustic reflection coefficient at the sample-transducer interface and the photoacoustic interaction inside the transparent sample, the density and compressibility of the sample can be probed simultaneously. Using an ad hoc signal analysis based on Hilbert and wavelet transforms, these quantities are measured accurately for a reference fluid. Similar analysis performed in a single vegetal cell also suggests high sensitivity to the state of the transducer-cell interface, and notably to the presence of the plasma membrane that encloses the cell vacuole.

  3. Interdependence of initial cell density, drug concentration and exposure time revealed by real-time impedance spectroscopic cytotoxicity assay

    DEFF Research Database (Denmark)

    Caviglia, Claudia; Zor, Kinga; Canepa, Silvia

    2015-01-01

    between the rate of cell death and the initial cell seeding density was found at 2.5 μM doxorubicin concentration, whereas this was not observed at 5 or 100 μM. By sensing the changes in the cell–substrate interaction using impedance spectroscopy under static conditions, the onset of cytotoxicity......We investigated the combined effect of the initial cell density (12 500, 35 000, 75 000, and 100 000 cells cm−2) and concentration of the anti-cancer drug doxorubicin on HeLa cells by performing timedependent cytotoxicity assays using real-time electrochemical impedance spectroscopy. A correlation...

  4. Genetic characterization of the hdrRM operon: a novel high-cell-density-responsive regulator in Streptococcus mutans.

    Science.gov (United States)

    Merritt, Justin; Zheng, Lanyan; Shi, Wenyuan; Qi, Fengxia

    2007-08-01

    Many species of bacteria can adhere to surfaces and grow as sessile communities. The continued accumulation of bacteria can eventually lead to the extremely high-cell-density environment characteristic of many biofilms or cell colonies. This is the normal habitat of the cariogenic species Streptococcus mutans, which normally resides in the high-cell-density, multispecies community commonly referred to as dental plaque. Previous work has demonstrated that the transcription of two separate bacteriocins can be activated by the high-cell-density conditions created through the centrifugation and incubation of cell pellets. In this study, we identified an uncharacterized two-gene operon that was induced >10-fold by conditions of high cell density. The genes of the operon encode a putative transcription regulator and a membrane protein, which were renamed as hdrR and hdrM, respectively. A transcription fusion to the hdrRM operon confirmed its induction by high cell density. Mutation of hdrM abolished bacteriocin production, greatly increased natural competence, reduced the growth rate, and severely affected biofilm formation. Interestingly, no obvious phenotypes were observed from a non-polar mutation of hdrR or mutations affecting the entire operon. These data suggest that the hdrRM operon may constitute a novel regulatory system responsible for mediating a cellular response to a high-cell-density environment.

  5. [Correlation with MRI features, cell density and the expression of immunohistochemistry of pure mucinous breast carcinoma].

    Science.gov (United States)

    Guo, Y; Kong, Q C; Zhu, Y Q; Liu, C L; He, H; Zhang, J E; Yang, R M; Jiang, X Q

    2017-05-09

    Objective: To explore the MRI features of the pure mucinous breast carcinoma(PMBC) and the correlation with cell density and the expression of immunohistochemistry. Methods: MRI features of 35 pure mucinous carcinomas were retrospectively analyzed from January 2011 to May 2016 in Guangdong General Hospital. MR images were reviewed for shape, margin, the signal intensity, enhancement patterns of tumors and diffusion weighted imaging (DWI) features and apparent diffusion coefficient (ADC) value. All the patients were detected by immunohistochemical staining with expression of ER, PR, CerbB-2, Ki-67 and Her-2. Correlations between MRI features of PMBC and cell density and the expression of immunohistochemistry were analyzed. Results: A total of 16 oval masses(16/35, 45.7%) and 10 round masses(10/35, 28.6%)were found in 35 PMBC with clear boundary(26/35, 74.3%) and lobulated shape(31/35, 88.6%). Very high signal intensity on T(2)-weighted images was found in 33 PMBC (33/35, 94.3%) and early enhancement rate was 115%±9% for PMBC. 28 PMBC demonstrated persistent enhancing pattern on time-signal intensity curve and 7 PMBC demonstrated plateau pattern.Mean ADC value was (1.91±0.06)×10(-3)mm(2)/s for PMBC. There was significant difference with early enhancement rate and ADC value between PMBC with more or less quantities of cellular mucin (P2, Her-2 and Ki-67 expression between PMBC with more or less quantities of cellular mucin (all P>0.05). Conclusions: PMBC has distinctive MRI features. The prognosis of PMBC is better from correlation between MRI features, cell density and the expression of immunohistochemistry.

  6. Improving accuracy of cell and chromophore concentration measurements using optical density.

    Science.gov (United States)

    Myers, John A; Curtis, Brandon S; Curtis, Wayne R

    2013-04-22

    UV-vis spectrophotometric optical density (OD) is the most commonly-used technique for estimating chromophore formation and cell concentration in liquid culture. OD wavelength is often chosen with little thought given to its effect on the quality of the measurement. Analysis of the contributions of absorption and scattering to the measured optical density provides a basis for understanding variability among spectrophotometers and enables a quantitative evaluation of the applicability of the Beer-Lambert law. This provides a rational approach for improving the accuracy of OD measurements used as a proxy for direct dry weight (DW), cell count, and pigment levels. For pigmented organisms, the choice of OD wavelength presents a tradeoff between the robustness and the sensitivity of the measurement. The OD at a robust wavelength is primarily the result of light scattering and does not vary with culture conditions; whereas, the OD at a sensitive wavelength is additionally dependent on light absorption by the organism's pigments. Suitably robust and sensitive wavelengths are identified for a wide range of organisms by comparing their spectra to the true absorption spectra of dyes. The relative scattering contribution can be reduced either by measurement at higher OD, or by the addition of bovine serum albumin. Reduction of scattering or correlation with off-peak light attenuation provides for more accurate assessment of chromophore levels within cells. Conversion factors between DW, OD, and colony-forming unit density are tabulated for 17 diverse organisms to illustrate the scope of variability of these correlations. Finally, an inexpensive short pathlength LED-based flow cell is demonstrated for the online monitoring of growth in a bioreactor at culture concentrations greater than 5 grams dry weight per liter which would otherwise require off-line dilutions to obtain non-saturated OD measurements. OD is most accurate as a time-saving proxy measurement for biomass

  7. Comparison of manual & automated analysis methods for corneal endothelial cell density measurements by specular microscopy.

    Science.gov (United States)

    Huang, Jianyan; Maram, Jyotsna; Tepelus, Tudor C; Modak, Cristina; Marion, Ken; Sadda, SriniVas R; Chopra, Vikas; Lee, Olivia L

    2017-08-07

    To determine the reliability of corneal endothelial cell density (ECD) obtained by automated specular microscopy versus that of validated manual methods and factors that predict such reliability. Sharp central images from 94 control and 106 glaucomatous eyes were captured with Konan specular microscope NSP-9900. All images were analyzed by trained graders using Konan CellChek Software, employing the fully- and semi-automated methods as well as Center Method. Images with low cell count (input cells number <100) and/or guttata were compared with the Center and Flex-Center Methods. ECDs were compared and absolute error was used to assess variation. The effect on ECD of age, cell count, cell size, and cell size variation was evaluated. No significant difference was observed between the Center and Flex-Center Methods in corneas with guttata (p=0.48) or low ECD (p=0.11). No difference (p=0.32) was observed in ECD of normal controls <40 yrs old between the fully-automated method and manual Center Method. However, in older controls and glaucomatous eyes, ECD was overestimated by the fully-automated method (p=0.034) and semi-automated method (p=0.025) as compared to manual method. Our findings show that automated analysis significantly overestimates ECD in the eyes with high polymegathism and/or large cell size, compared to the manual method. Therefore, we discourage reliance upon the fully-automated method alone to perform specular microscopy analysis, particularly if an accurate ECD value is imperative. Copyright © 2017. Published by Elsevier España, S.L.U.

  8. Cultivation of microalgae in industrial wastewaters

    DEFF Research Database (Denmark)

    van Wagenen, Jonathan Myerson

    that has many potential uses. Unfortunately, the current high costs of cultivation have limited the development and exploitation of such systems, resulting in only a few full-scale algae wastewater treatment installations and a small industry based mostly around food and pigments. This thesis contributes...... to autotrophic controls. Industrial wastewater was used as cultivation medium of Chlorella sorokiniana. The culture was able to grow at high rates upto a density of 4 g L-1. The deceleration-stat technique was used to create a series of pseudo-steady states to give information about the expected results...... to a growing body of knowledge with the aim to make algae cultivation viable for the production of sustainable products. Specific contributions include: improvement in the methods of screening the growth potential of different microalgae species; identification of an industrial wastewater that allows good...

  9. Goblet cell density estimate differences in impression cytology samples varies with different magnification of images.

    Science.gov (United States)

    Doughty, Michael J

    2017-12-04

    To assess the impact of using different microscope magnifications for the goblet cell density (GCD) estimates from conjunctival impression cytology (CIC) samples from healthy individuals METHODS: In a prospective study, CIC specimens were collected from the superior bulbar conjunctiva (12 o'clock, 5mm from limbus) of 20 adult subjects (average age 22 years) onto Millicell-CM membranes and Giemsa stained. A region from each CIC filter containing reasonably high numbers of goblet cells was imaged by light microscopy at a final magnification of 400X and then the same region assessed at 200X and then 100X. The images were enlarged, the goblet cells marked and counted and GCD values/sq mm calculated. The mean GCD estimates at 400X magnification, 200X and 100X were 644±180, 405±72 and 365±81 cells/sq mm respectively, and these values were statistically different (pmicroscope field (HPF) that appears to include a moderate number of goblet cells will have a probability (by at least 20:1) that the GCD estimates will likely be higher compared to those at 200X or 100X, and the probability for higher GCD values is at least 15:1 comparing assessments made at 200X to 100X. Investigators should use only one magnification, with that of a medium power field (200X final magnification) likely being the most useful. Copyright © 2017 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

  10. High-energy-density hydrogen-halogen fuel cells for advanced military applications

    Science.gov (United States)

    Balko, E. N.; McElroy, J. F.

    It is pointed out that hydrogen-halogen fuel cell systems are particularly suited for an employment as ground power sources for military applications. The large cell potential and reversible characteristics of the H2/Cl2 and H2/Br2 couples permit high energy storage density and efficient energy conversion. When used as flow batteries, the fluid nature of the reactants in the hydrogen-halogen systems has several advantages over power sources which involve solid phases. Very deep discharge is possible without degradation of subsequent performance, and energy storage capacity is limited only by the external reactant storage volume. Very rapid chemical recharging is possible through replenishment of the reactant supply. A number of H2/Cl2 and H2/Br2 fuel cell systems have been studied. These systems use the same solid polymer electrolyte (SPE) cell technology originally developed for H2/O2 fuel cells. The results of the investigation are illustrated with the aid of a number of graphs.

  11. Bone mineral density, growth, pubertal development and other parameters in Brazilian children and young adults with sickle cell anaemia

    NARCIS (Netherlands)

    Meeuwes, M.; Souza de Carvalho, T. F.; Cipolotti, R.; Gurgel, R. Q.; Ferrão, T. O.; Peters, M.; Agyemang, C.

    2013-01-01

    To evaluate the occurrence of low bone mineral density (BMD) and its relationship with clinical and laboratorial characteristics in children and young adults with sickle cell anaemia living in Northeast-Brazil, and to assess the role of radiography in diagnosing low BMD. Bone mineral density of

  12. An escort for GPCRs: implications for regulation of receptor density at the cell surface.

    Science.gov (United States)

    Achour, Lamia; Labbé-Jullié, Catherine; Scott, Mark G H; Marullo, Stefano

    2008-10-01

    G-protein-coupled receptors (GPCRs) are dynamically regulated by various mechanisms that tune their response to external stimuli. Modulation of their plasma membrane density, via trafficking between subcellular compartments, constitutes an important process in this context. Substantial information has been accumulated on cellular pathways that remove GPCRs from the cell surface for subsequent degradation or recycling. In comparison, much less is known about the mechanisms controlling trafficking of neo-synthesized GPCRs from intracellular compartments to the cell surface. Although GPCR export to the plasma membrane is commonly considered to mostly implicate the default, unregulated secretory pathway, an increasing number of observations indicate that trafficking to the plasma membrane from the endoplasmic reticulum might be tightly regulated and involve specific protein partners. Moreover, a new paradigm is emerging in some cellular contexts, in which stocks of functional receptors retained within intracellular compartments can be rapidly mobilized to the plasma membrane to maintain sustained physiological responsiveness.

  13. Mesenchymal stem cell attachment to peptide density gradients on porous silicon generated by electrografting

    Energy Technology Data Exchange (ETDEWEB)

    Clements, Lauren R. [School of Chemical and Physical Sciences, Flinders University, Bedford Park, SA (Australia); CSIRO Molecular and Health Technologies, Bayview Avenue, Clayton, VIC (Australia); Wang, Peng-Yuan; Tsai, Wei-Bor [Department of Chemical Engineering, National Taiwan University, Taipei (China); Harding, Frances; Voelcker, Nicolas H. [School of Chemical and Physical Sciences, Flinders University, Bedford Park, SA (Australia); Thissen, Helmut [CSIRO Molecular and Health Technologies, Bayview Avenue, Clayton, VIC (Australia)

    2011-06-15

    Chemical gradients of ethyl-6-bromohexanoate (EBH) were generated over porous silicon (pSi) substrates via electrochemical attachment. Immobilised ester moieties were hydrolysed and activated to produce a gradient of functional carboxylic acid groups. After subsequent immobilisation of cyclic RGD (cRGD), the surfaces were used to screen the extent of rat mesenchymal stem cell (MSC) attachment. Mapping of surface chemistry was carried out by means of infrared microscopy and X-ray photoelectron spectroscopy (XPS). MSC culture studies showed that short-term cell attachment responded to the cRGD density present on the gradient surface. (Copyright copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  14. Fast wave experiments in LAPD: RF sheaths, convective cells and density modifications

    Science.gov (United States)

    Carter, T. A.; van Compernolle, B.; Martin, M.; Gekelman, W.; Pribyl, P.; van Eester, D.; Crombe, K.; Perkins, R.; Lau, C.; Martin, E.; Caughman, J.; Tripathi, S. K. P.; Vincena, S.

    2017-10-01

    An overview is presented of recent work on ICRF physics at the Large Plasma Device (LAPD) at UCLA. The LAPD has typical plasma parameters ne 1012 -1013 cm-3, Te 1 - 10 eV and B 1000 G. A new high-power ( 150 kW) RF system and fast wave antenna have been developed for LAPD. The source runs at a frequency of 2.4 MHz, corresponding to 1 - 7fci , depending on plasma parameters. Evidence of rectified RF sheaths is seen in large increases ( 10Te) in the plasma potential on field lines connected to the antenna. The rectified potential scales linearly with antenna current. The rectified RF sheaths set up convective cells of local E × B flows, measured indirectly by potential measurements, and measured directly with Mach probes. At high antenna powers substantial modifications of the density profile were observed. The plasma density profile initially exhibits transient low frequency oscillations (10 kHz). The amplitude of the fast wave fields in the core plasma is modulated at the same low frequency, suggesting fast wave coupling is affected by the density rearrangement. Work performed at the Basic Plasma Science Facility, supported jointly by the National Science Foundation and the Department of Energy.

  15. Ceruloplasmin enhances smooth muscle cell- and endothelial cell-mediated low density lipoprotein oxidation by a superoxide-dependent mechanism

    Science.gov (United States)

    Mukhopadhyay, C. K.; Ehrenwald, E.; Fox, P. L.

    1996-01-01

    Cultured vascular smooth muscle cells (SMC) and endothelial cells (EC) stimulate low density lipoprotein (LDL) oxidation by free radical-mediated, transition metal-dependent mechanisms. The physiological source(s) of metal ions is not known; however, purified ceruloplasmin, a plasma protein containing 7 coppers, oxidizes LDL in vitro. We now show that ceruloplasmin also increases LDL oxidation by vascular cells. In metal ion-free medium, human ceruloplasmin increased bovine aortic SMC- and EC-mediated LDL oxidation by up to 30- and 15-fold, respectively. The maximal response was at 100-300 microg ceruloplasmin/ml, a level at or below the unevoked physiological plasma concentration. Oxidant activity was dependent on protein structure as a specific proteolytic cleavage or removal of one of the seven ceruloplasmin copper atoms inhibited activity. Three lines of evidence indicated a critical role for cellular superoxide (O2.) in ceruloplasmin-stimulated oxidation. First, the rate of production of O2. by cells correlated with their rates of LDL oxidation. Second, superoxide dismutase effectively blocked ceruloplasmin-stimulated oxidation by both cell types. Finally, O2. production by SMC quantitatively accounted for the observed rate of LDL oxidation. To show this, the course of O2. production by SMC was simulated by repeated addition of xanthine and xanthine oxidase to culture medium under cell-free conditions. Neither ceruloplasmin nor O2. alone increased LDL oxidation, but together they completely reconstituted the oxidation rate of ceruloplasmin-stimulated SMC. These results are the first to show that ceruloplasmin stimulates EC- and SMC-mediated oxidation of LDL and that cell-derived O2. accounts quantitatively for metal-dependent, free radical-initiated oxidation of LDL by these cells.

  16. Corneal endothelial cell density and morphology and central corneal thickness in Guangxi Maonan and Han adolescent students of China

    Directory of Open Access Journals (Sweden)

    Hao Liang

    2015-06-01

    Full Text Available AIM:To investigate the corneal endothelial cell density and morphology and central corneal thickness in the Guangxi Maonan and Han adolescent students of China.METHODS:Noncontact specular microscope (Topcon SP3000P, Tokyo, Japan was performed in 133 adolescent students of Maonan nationality (M:F 54:79 and 105 adolescent students of Han nationality (M:F 50:55, 5 to 20y of age, who were randomly selected from 3 schools in Huanjiang Maonan Autonomous County of Guangxi Zhuang Autonomous Region of China. Parameters studied included endothelial cell density, mean cell area, coefficient of variation in cell size, percentage hexagonality and central corneal thickness.RESULTS:Endothelial cell density, mean cell area, coefficient of variation in cell size, percentage hexagonality and central corneal thickness in the study population were (2969.50±253.93 cells/mm2, (339.23±29.44 µm2, (29.96±4.07 %, (64.58±9.41 % and (523.71±32.82 µm in Maonan and (2998.26±262.65 cells/mm2, (336.11±30.07 µm2, (29.89±5.03 %, (64.91±11.64 % and (524.39±33.15 µm in Han, respectively. No significant differences were observed in endothelial cell density, mean cell area, coefficient of variation in cell size, percentage hexagonality and central corneal thickness between Maonan and Han (P=0.615, 0.659, 0.528, 0.551, 0.999. In Maonan and Han, we found age was negatively correlated with endothelial cell density and percentage hexagonality and positively correlated with mean cell area and coefficient of variation in cell size. Negative correlation was also found between central corneal thickness and age in Han, whereas no correlation was found in Maonan.CONCLUSION:There were no differences between Maonan and Han in corneal endothelial cell density and morphology and central corneal thickness. In these two nationalities, there were statistically significant decrease in endothelial cell density and percentage hexagonality with increasing age and statistically significant

  17. [Using Excess Activated Sludge Treated 4-Chlorophenol Contained Waste Water to Cultivate Chlorella vulgaris].

    Science.gov (United States)

    Wang, Lu; Chen, Xiu-rong; Yan, Long; He, Yi-xuan; Shi, Zhen-dong

    2015-04-01

    Using different rations of sludge extracts and supernate from 4-Chlorophenol (4-CP) simulated wastewater's excess sludge after centrifugation to cultivate the Chlorella vulgaris to achieve the goal of excess sludge utilization together with chlorella cultivating. The experiments were performed in 500 mL flasks with different rations of sludge extracts & BG-11 and supernate & BG-11 in a light growth chamber respectively. Number of algal cells, Chlorophyll, enzyme activity, oil and water total nitrogen (TN), total phosphorus (TP), total organic carbon (TOC), toxicity index were investigated. Result showed that the nutrition supplies and toxicity in the excess sludge were removed efficiently via Chlorella vulgaris, the removal rates of TN and TP were at least 40% and 90% respectively; After 10 days cultivation, the density growth of 50% sludge extracts was 20 times higher of the beginning while its chlorophyll content was lower than that of the blank group. Sludge extracts could promote the proliferation of algae, but were not conducive to the synthesis of chlorophyll. The quantity of SOD in per cell showed Chlorella vulgaris gave a positive response via stimulation from toxicant in sludge extracts and supernate. The best time for collecting chlorella vulgaris was the fifth day of cultivation, taking neutral oil accumulation as the evaluating indicator for its utilization combined with the removal of supplies and toxicity.

  18. Higher-Density Culture in Human Embryonic Stem Cells Results in DNA Damage and Genome Instability

    Science.gov (United States)

    Jacobs, Kurt; Zambelli, Filippo; Mertzanidou, Afroditi; Smolders, Ilse; Geens, Mieke; Nguyen, Ha Thi; Barbé, Lise; Sermon, Karen; Spits, Claudia

    2016-01-01

    Summary Human embryonic stem cells (hESC) show great promise for clinical and research applications, but their well-known proneness to genomic instability hampers the development to their full potential. Here, we demonstrate that medium acidification linked to culture density is the main cause of DNA damage and genomic alterations in hESC grown on feeder layers, and this even in the short time span of a single passage. In line with this, we show that increasing the frequency of the medium refreshments minimizes the levels of DNA damage and genetic instability. Also, we show that cells cultured on laminin-521 do not present this increase in DNA damage when grown at high density, although the (long-term) impact on their genomic stability remains to be elucidated. Our results explain the high levels of genome instability observed over the years by many laboratories worldwide, and show that the development of optimal culture conditions is key to solving this problem. PMID:26923824

  19. Durability of Low Platinum Fuel Cells Operating at High Power Density

    Energy Technology Data Exchange (ETDEWEB)

    Polevaya, Olga [Nuvera Fuel Cells Inc.; Blanchet, Scott [Nuvera Fuel Cells Inc.; Ahluwalia, Rajesh [Argonne National Lab; Borup, Rod [Los-Alamos National Lab; Mukundan, Rangachary [Los-Alamos National Lab

    2014-03-19

    Understanding and improving the durability of cost-competitive fuel cell stacks is imperative to successful deployment of the technology. Stacks will need to operate well beyond today’s state-of-the-art rated power density with very low platinum loading in order to achieve the cost targets set forth by DOE ($15/kW) and ultimately be competitive with incumbent technologies. An accelerated cost-reduction path presented by Nuvera focused on substantially increasing power density to address non-PGM material costs as well as platinum. The study developed a practical understanding of the degradation mechanisms impacting durability of fuel cells with low platinum loading (≤0.2mg/cm2) operating at high power density (≥1.0W/cm2) and worked out approaches for improving the durability of low-loaded, high-power stack designs. Of specific interest is the impact of combining low platinum loading with high power density operation, as this offers the best chance of achieving long-term cost targets. A design-of-experiments approach was utilized to reveal and quantify the sensitivity of durability-critical material properties to high current density at two levels of platinum loading (the more conventional 0.45 mgPt.cm–1 and the much lower 0.2 mgPt.cm–2) across several cell architectures. We studied the relevance of selected component accelerated stress tests (AST) to fuel cell operation in power producing mode. New stress tests (NST) were designed to investigate the sensitivity to the addition of electrical current on the ASTs, along with combined humidity and load cycles and, eventually, relate to the combined city/highway drive cycle. Changes in the cathode electrochemical surface area (ECSA) and average oxygen partial pressure on the catalyst layer with aging under AST and NST protocols were compared based on the number of completed cycles. Studies showed elevated sensitivity of Pt growth to the potential limits and the initial particle size distribution. The ECSA loss

  20. Kinetic modeling of rhamnolipid production by Pseudomonas aeruginosa PAO1 including cell density-dependent regulation.

    Science.gov (United States)

    Henkel, Marius; Schmidberger, Anke; Vogelbacher, Markus; Kühnert, Christian; Beuker, Janina; Bernard, Thomas; Schwartz, Thomas; Syldatk, Christoph; Hausmann, Rudolf

    2014-08-01

    The production of rhamnolipid biosurfactants by Pseudomonas aeruginosa is under complex control of a quorum sensing-dependent regulatory network. Due to a lack of understanding of the kinetics applicable to the process and relevant interrelations of variables, current processes for rhamnolipid production are based on heuristic approaches. To systematically establish a knowledge-based process for rhamnolipid production, a deeper understanding of the time-course and coupling of process variables is required. By combining reaction kinetics, stoichiometry, and experimental data, a process model for rhamnolipid production with P. aeruginosa PAO1 on sunflower oil was developed as a system of coupled ordinary differential equations (ODEs). In addition, cell density-based quorum sensing dynamics were included in the model. The model comprises a total of 36 parameters, 14 of which are yield coefficients and 7 of which are substrate affinity and inhibition constants. Of all 36 parameters, 30 were derived from dedicated experimental results, literature, and databases and 6 of them were used as fitting parameters. The model is able to describe data on biomass growth, substrates, and products obtained from a reference batch process and other validation scenarios. The model presented describes the time-course and interrelation of biomass, relevant substrates, and products on a process level while including a kinetic representation of cell density-dependent regulatory mechanisms.

  1. The three-dimensional cultivation of the carcinoma cell line HepG2 in a perfused chip system leads to a more differentiated phenotype of the cells compared to monolayer culture

    Energy Technology Data Exchange (ETDEWEB)

    Altmann, B; Giselbrecht, S; Weibezahn, K-F; Welle, A; Gottwald, E [Forschungszentrum Karlsruhe, Institute for Biological Interfaces, 76344 Eggenstein-Leopoldshafen (Germany)], E-mail: eric.gottwald@ibg.fzk.de

    2008-09-01

    We describe a polymer chip with a grid-like architecture that it is intended for the three-dimensional cultivation of cells with an active nutrient and gas supply. The chip is typically made from polymethyl methacrylate or polycarbonate but can also be manufactured from biodegradable polymers, such as poly(lactic-co-glycolic acid). Different designs of the chip can be realized. In this study, we evaluated a chip with 506 microcontainers of the size of 300 x 300 x 300 {mu}m that are capable of housing up to 6 million cells, and its suitability as a tissue-specific culture system for the carcinoma cell line HepG2 instead of primary liver cells. Related to an earlier study, where we could show the principal suitability of the system for rat primary cells, we here investigated the system's suitability for the human carcinoma cell line HepG2. The carcinoma cells were used in two different types of chip-containing bioreactors. By confocal laser scanning microscopy, we could show that cellular integrity in the chip culture was maintained and that there were no signs of apoptosis as confirmed by the absence of K18 fragmentation. Gene expression analysis of some liver-specific genes revealed a significantly higher expression of the phase II metabolism genes uridine-diphosphate- glucosyl-transferase (UGT1A1) and glutathione-S-transferase (GST{pi}1) as a marker. Therefore, we conclude that by using a three-dimensional instead of a conventional monolayer culture system, hepatocellular carcinoma cells display a phenotype that resembles more closely the tissue of origin.

  2. Tailoring the rate-sensitivity of low density polyurea foams through cell wall aperture size

    Science.gov (United States)

    Ramirez, B. J.; Kingstedt, O. T.; Crum, R.; Gamez, C.; Gupta, V.

    2017-06-01

    The plateau stress and energy absorption of low density (≤300 kg/m3) polyurea (PU) foams and expanded polystyrene (EPS) were measured at deformation rates ranging from 0.004 s-1 to 5000 s-1. Low (≤10-1 s-1) strain rate testing was performed using an Instron load frame, intermediate (101-102 s-1) strain rates using a drop-weight impact tower, and high (≥103 s-1) strain rate conditions using a modified split-Hopkinson pressure bar. The plateau stress and energy absorption of low density PU foams exhibit a strong rate dependence across all deformation rates. This result has been previously unreported for low density polymer foams under low and intermediate strain rates. The strain rate sensitivity of PU foams was found to be strongly dependent on cell size for low strain rates and cell wall aperture size for intermediate and high strain rates. EPS type foam, however, remained nearly insensitive to strain rate. At low and intermediate strain rates, the plastic crushing in the EPS and the high plateau stress yield a much higher energy absorption capability than the viscoelastic dissipation in the PU foams. However, PU foams were found to display similar energy absorption properties as EPS based foams under high strain rates. Thus, controlling the strain rate sensitivity of PU foams through aperture diameter can lead to an increase in energy absorption properties at high strain rates, while simultaneously maintaining the peak stress below certain injury thresholds. Additionally, unlike EPS, which undergo plastic crushing after first impact, flexible polyurea foams will recover fully after each impact and thus will have multiple hit capabilities. This will allow these materials to have a wide range of applications, in advance body armors and protective headgears to use in low-cost protection systems for a wide range of military platforms, civilian, and space applications.

  3. [A Modified Procedure to Isolate Synchronous Cells from Yeasts with Continuous Percoll Density Gradient and Their Raman Discrimination].

    Science.gov (United States)

    Huang, Shu-shi; Lai, Jun-zhuo; Lu, Ming-qian; Cheng, Qin; Liao, Wei; Chen, Li-mei

    2015-08-01

    A modified procedure of Percoll density gradient centrifugation was developed to isolate and fractionate synchronous cells from stationary phase (sp) cultures of different yeast strains, as well as Raman spectra discrimination of single yeast cells was reported. About 1.75 mL Percoll solution in 2 mL polypropylene centrifugal tube was centrifuged at 19,320 g, 20 °C with an angle rotor for 15 min to form continuous densities gradient (1.00~1.31 g · mL(-1)), approximately 100 μL sample was overlaid onto the preformed continuous density gradient carefully, subsequently, centrifuged at 400 g for 60 min in a tabletop centrifuge equipped with a angle rotor at 25 °C. Yeast samples could be observed that the suspensions were separated into two cell fractions obviously. Both fractions of different yeast strains were respectively determined by differential interference contrast (DIC), phase contrast microscope and synchronous culture to distinguish their morphological and growth trait. The results showed that the lower fraction cells were unbudded, mostly unicellular, highly refractive, homogeneous and uniform in size, and represented growth characteristic synchronously; Their protoplasm had relatively high density, and contained significant concentrations of glycogen; all of which were accordant with description of quiescent yeast cells and G0 cells in previously published paper. It was shown that lower fraction was quiescent cells, synchronous G0 cells as well. A Raman tweezers setup was used to investigate the differences between two fractions, G0 cells and non G0 cells, at a single cell level. The result showed that both G0 cells and the non G0 cells had the same characteristic peaks corresponding biological macromolecules including proteins, carbohydrates and nucleic acids, but all characteristic peak intensities of G0 cells were higher than that of non G0 cells, implied that the macromolecular substance content of G0 cells was more higher. Principal component

  4. Poly(3-Hydroxybutyrate) Production from Glycerol by Zobellella denitrificans MW1 via High-Cell-Density Fed-Batch Fermentation and Simplified Solvent Extraction▿

    Science.gov (United States)

    Ibrahim, Mohammad H. A.; Steinbüchel, Alexander

    2009-01-01

    Industrial production of biodegradable polyesters such as polyhydroxyalkanoates is hampered by high production costs, among which the costs for substrates and for downstream processing represent the main obstacles. Inexpensive fermentable raw materials such as crude glycerol, an abundant by-product of the biodiesel industry, have emerged to be promising carbon sources for industrial fermentations. In this study, Zobellella denitrificans MW1, a recently isolated bacterium, was used for the production of poly(3-hydroxybutyrate) (PHB) from glycerol as the sole carbon source. Pilot-scale fermentations (42-liter scale) were conducted to scale up the high PHB accumulation capability of this strain. By fed-batch cultivation, at first a relatively high cell density (29.9 ± 1.3 g/liter) was obtained during only a short fermentation period (24 h). However, the PHB content was relatively low (31.0% ± 4.2% [wt/wt]). Afterwards, much higher concentrations of PHB (up to 54.3 ± 7.9 g/liter) and higher cell densities (up to 81.2 ± 2.5 g/liter) were obtained by further fed-batch optimization in the presence of 20 g/liter NaCl, with optimized feeding of glycerol and ammonia to support both cell growth and polymer accumulation over a period of 50 h. A high specific growth rate (0.422/h) and a short doubling time (1.64 h) were attained. The maximum PHB content obtained was 66.9% ± 7.6% of cell dry weight, and the maximum polymer productivity and substrate yield coefficient were 1.09 ± 0.16 g/liter/h and 0.25 ± 0.04 g PHB/g glycerol, respectively. Furthermore, a simple organic solvent extraction process was employed for PHB recovery during downstream processing: self-flotation of cell debris after extraction of PHB with chloroform allowed a convenient separation of a clear PHB-solvent solution from the cells. Maximum PHB recovery (85.0% ± 0.10% [wt/wt]) was reached after 72 h of extraction with chloroform at 30°C, with a polymer purity of 98.3% ± 1.3%. PMID:19666728

  5. Low Mast Cell Density Predicts Poor Prognosis in Oral Squamous Cell Carcinoma and Reduces Survival in Head and Neck Squamous Cell Carcinoma.

    Science.gov (United States)

    Attramadal, Cecilie Gjøvaag; Kumar, Sheeba; Gao, Jian; Boysen, Morten Ebbe; Halstensen, Trond Sundby; Bryne, Magne

    2016-10-01

    The cellular composition of the tumor microenvironment (TME) at the invading front of oral squamous cell carcinomas (OSCCs) may reflect biologically important cancer features and host responses, and thus be related to disease progression. The TME density of mast cells (MCs), macrophages, cancer-associated fibroblasts (CAFs) and endothelial cells were quantified at the invasive front and analyzed regarding their relation to disease recurrence in patients with small T1/2N0M0 OSCCs. mRNA for MC-specific proteins were analyzed in a second patient cohort with head and neck squamous cell carcinoma (HNSCC). Samples from 62 patients with T1/2N0M0 OSCC were immunohistochemically stained and scored for the cellular expression of mast/stem cell growth factor receptor (c-KIT) (MCs), CD68 (macrophages), α-smooth muscle actin (α-SMA) (CAFs) and CD31 (endothelial cells) and this was analyzed according to disease recurrence. Data from The Cancer Genome Atlas database were used to examine mRNA expression profiles and clinical data of patients with 399 HNSCC. Increased MC density at the invasive front was significantly associated with reduced disease recurrence, as none of the patients with high MC density experienced relapse. Moreover, increased expression of mRNA for MC specific markers as c-KIT, and α-, β-, and δ-tryptases and the MC-stimulating factor, stem cell factor (SCF), was significantly associated with good prognosis in patients with HNSCC. Decreased MC density at the invasive front may reflect tumor biology related to disease progression and prognosis. Counting MCs seems to be an easy and practical tool, that could be utilized for prognostic evaluation. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  6. Enhanced photocurrent density in graphene/Si based solar cell (GSSC) by optimizing active layer thickness

    Energy Technology Data Exchange (ETDEWEB)

    Rosikhin, Ahmad, E-mail: a.rosikhin86@yahoo.co.id; Hidayat, Aulia Fikri; Syuhada, Ibnu; Winata, Toto, E-mail: toto@fi.itb.ac.id [Department of physics, physics of electronic materials research division Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung Jl. Ganesha 10, Bandung 40132, Jawa Barat – Indonesia (Indonesia)

    2015-12-29

    Thickness dependent photocurrent density in active layer of graphene/Si based solar cell has been investigated via analytical – simulation study. This report is a preliminary comparison of experimental and analytical investigation of graphene/Si based solar cell. Graphene sheet was interfaced with Si thin film forming heterojunction solar cell that was treated as a device model for photocurrent generator. Such current can be enhanced by optimizing active layer thickness and involving metal oxide as supporting layer to shift photons absorption. In this case there are two type of devices model with and without TiO{sub 2} in which the silicon thickness varied at 20 – 100 nm. All of them have examined and also compared with each other to obtain an optimum value. From this calculation it found that generated currents almost linear with thickness but there are saturated conditions that no more enhancements will be achieved. Furthermore TiO{sub 2} layer is effectively increases photon absorption but reducing device stability, maximum current is fluctuates enough. This may caused by the disturbance of excitons diffusion and resistivity inside each layer. Finally by controlling active layer thickness, it is quite useful to estimate optimization in order to develop the next solar cell devices.

  7. Enhanced photocurrent density in graphene/Si based solar cell (GSSC) by optimizing active layer thickness

    Science.gov (United States)

    Rosikhin, Ahmad; Hidayat, Aulia Fikri; Syuhada, Ibnu; Winata, Toto

    2015-12-01

    Thickness dependent photocurrent density in active layer of graphene/Si based solar cell has been investigated via analytical - simulation study. This report is a preliminary comparison of experimental and analytical investigation of graphene/Si based solar cell. Graphene sheet was interfaced with Si thin film forming heterojunction solar cell that was treated as a device model for photocurrent generator. Such current can be enhanced by optimizing active layer thickness and involving metal oxide as supporting layer to shift photons absorption. In this case there are two type of devices model with and without TiO2 in which the silicon thickness varied at 20 - 100 nm. All of them have examined and also compared with each other to obtain an optimum value. From this calculation it found that generated currents almost linear with thickness but there are saturated conditions that no more enhancements will be achieved. Furthermore TiO2 layer is effectively increases photon absorption but reducing device stability, maximum current is fluctuates enough. This may caused by the disturbance of excitons diffusion and resistivity inside each layer. Finally by controlling active layer thickness, it is quite useful to estimate optimization in order to develop the next solar cell devices.

  8. Metal based gas diffusion layers for enhanced fuel cell performance at high current densities

    Science.gov (United States)

    Hussain, Nabeel; Van Steen, Eric; Tanaka, Shiro; Levecque, Pieter

    2017-01-01

    The gas diffusion layer strongly influences the performance and durability of polymer electrolyte fuel cells. A major drawback of current carbon fiber based GDLs is the non-controlled variation in porosity resulting in a random micro-structure. Moreover, when subjected to compression these materials show significant reduction in porosity and permeability leading to water management problems and mass transfer losses within the fuel cell. This study investigated the use of uniform perforated metal sheets as GDLs in conjunction with microchannel flowfields. A metal sheet design with a pitch of 110 μm and a hole diameter of 60 μm in combination with an MPL showed superior performance in the high current density region compared to a commercially available carbon paper based GDL in a single cell environment. Fuel cell testing with different oxidants (air, heliox and oxygen) indicate that the metal sheet offers both superior diffusion and reduced flooding in comparison to the carbon based GDL. The presence of the MPL has been found to be critical to the functionality of the metal sheet suggesting that the MPL design may represent an important optimisation parameter for further improvements in performance.

  9. Common pediatric cerebellar tumors: correlation between cell densities and apparent diffusion coefficient metrics.

    Science.gov (United States)

    Koral, Korgün; Mathis, Derek; Gimi, Barjor; Gargan, Lynn; Weprin, Bradley; Bowers, Daniel C; Margraf, Linda

    2013-08-01

    To test whether there is correlation between cell densities and apparent diffusion coefficient (ADC) metrics of common pediatric cerebellar tumors. This study was reviewed for issues of patient safety and confidentiality and was approved by the Institutional Review Board of the University of Texas Southwestern Medical Center and was compliant with HIPAA. The need for informed consent was waived. Ninety-five patients who had preoperative magnetic resonance imaging and surgical pathologic findings available between January 2003 and June 2011 were included. There were 37 pilocytic astrocytomas, 34 medulloblastomas (23 classic, eight desmoplastic-nodular, two large cell, one anaplastic), 17 ependymomas (13 World Health Organization [WHO] grade II, four WHO grade III), and seven atypical teratoid rhabdoid tumors. ADCs of solid tumor components and normal cerebellum were measured. Tumor-to-normal brain ADC ratios (hereafter, ADC ratio) were calculated. The medulloblastomas and ependymomas were subcategorized according to the latest WHO classification, and tumor cellularity was calculated. Correlation was sought between cell densities and mean tumor ADCs, minimum tumor ADCs, and ADC ratio. When all tumors were considered together, negative correlation was found between cellularity and mean tumor ADCs (ρ = -0.737, P correlation between cellularity and ADC ratio. Negative correlation was found between cellularity and minimum tumor ADC in atypical teratoid rhabdoid tumors (ρ = -0.786, P correlation was found between cellularity and mean tumor ADC and ADC ratio. There was no correlation between the ADC metrics and cellularity of the pilocytic astrocytomas, medulloblastomas, and ependymomas. Negative correlation was found between cellularity and ADC metrics of common pediatric cerebellar tumors. Although ADC metrics are useful in the preoperative diagnosis of common pediatric cerebellar tumors and this utility is generally attributed to differences in cellularity of tumors

  10. Transition in nori cultivation

    DEFF Research Database (Denmark)

    Delaney, Alyne

    2011-01-01

    of social and environmental sustainability, we must understand both society and cultural institutions. With this in mind, this article focuses on the division of labor among cultivators, particularly along gender lines and the impacts, on a cultural level, of technological change on nori production...

  11. Cultivating the Grapevine.

    Science.gov (United States)

    Rauschenberg, Gretchen

    1988-01-01

    Although administrators may view grapevines as threats to established leadership, informal communication is necessary to keep an organization functioning smoothly. No one can completely control the grapevine or its accuracy, but astute administrators can learn to cultivate and use informal communication systems wisely. (MLH)

  12. High Cell Density Process for Constitutive Production of a Recombinant Phytase in Thermotolerant Methylotrophic Yeast Ogataea thermomethanolica Using Table Sugar as Carbon Source.

    Science.gov (United States)

    Charoenrat, Theppanya; Antimanon, Sompot; Kocharin, Kanokarn; Tanapongpipat, Sutipa; Roongsawang, Niran

    2016-12-01

    The yeast Ogataea thermomethanolica has recently emerged as a potential host for heterologous protein expression at elevated temperature. To evaluate the feasibility of O. thermomethanolica as heterologous host in large-scale fermentation, constitutive production of fungal phytase was investigated in fed-batch fermentation. The effect of different temperatures, substrate feeding strategies, and carbon sources on phytase production was investigated. It was found that O. thermomethanolica can grow in the temperature up to 40 °C and optimal at 34 °C. However, the maximum phytase production was observed at 30 °C and slightly decreased at 34 °C. The DOT stat control was the most efficient feeding strategy to obtain high cell density and avoid by-product formation. The table sugar can be used as an alternative substrate for phytase production in O. thermomethanolica. The highest phytase activity (134 U/mL) was obtained from table sugar at 34 °C which was 20-fold higher than batch culture (5.7 U/mL). At a higher cultivation temperature of 38 °C, table sugar can be used as a low-cost substrate for the production of phytase which was expressed with an acceptable yield (85 U/mL). Lastly, the results from this study reveal the industrial favorable benefits of employing O. thermomethanolica as a host for heterologous protein production.

  13. Cell Density Effects of Frog Skin Bacteria on Their Capacity to Inhibit Growth of the Chytrid Fungus, Batrachochytrium dendrobatidis.

    Science.gov (United States)

    Yasumiba, Kiyomi; Bell, Sara; Alford, Ross

    2016-01-01

    Bacterial symbionts on frog skin can reduce the growth of the chytrid fungus Batrachochytrium dendrobatidis (Bd) through production of inhibitory metabolites. Bacteria can be effective at increasing the resistance of amphibians to chytridiomycosis when added to amphibian skin, and isolates can be screened for production of metabolites that inhibit Bd growth in vitro. However, some bacteria use density-dependent mechanism such as quorum sensing to regulate metabolite production. It is therefore important to consider cell density effects when evaluating bacteria as possible candidates for bioaugmentation. The aim of our study was to evaluate how the density of cutaneous bacteria affects their inhibition of Bd growth in vitro. We sampled cutaneous bacteria isolated from three frog species in the tropical rainforests of northern Queensland, Australia, and selected ten isolates that were inhibitory to Bd in standardised pilot trials. We grew each isolate in liquid culture at a range of initial dilutions, sub-sampled each dilution at a series of times during the first 48 h of growth and measured spectrophotometric absorbance values, cell counts and Bd-inhibitory activity of cell-free supernatants at each time point. The challenge assay results clearly demonstrated that the inhibitory effects of most isolates were density dependent, with relatively low variation among isolates in the minimum cell density needed to inhibit Bd growth. We suggest the use of minimum cell densities and fast-growing candidate isolates to maximise bioaugmentation efforts.

  14. The Effect of Cultivated Wild Ginseng Extract on Preadipocyte Proliferation

    Directory of Open Access Journals (Sweden)

    Byoung-Woo Kim

    2007-12-01

    Full Text Available Objectives : The purpose of this study is to investigate the effects of cultivated wild ginseng extract on primary cultured preadipocyte and adipocytes. Methods : Diminish preadipocyte proliferation does primary role to reduce obesity. So, preadipocytes and adipocytes were performed on cell cultures with using Sprague-Dawley rats and treated with 0.01-1mg/㎖ cultivated wild ginseng extract. Result : At all concentrations, cultivated wild ginseng extract wasn't show the suppress proliferation of preadipocytes significantly and failed to show effects on decomposition of adipocytes except high dosage. Conclusion : Based on these findings, cultivated wild ginseng is not a suitable choice for the treatment of localized obesity.

  15. In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

    Directory of Open Access Journals (Sweden)

    T.I. Wodewotzky

    2012-12-01

    Full Text Available Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium.

  16. In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

    Energy Technology Data Exchange (ETDEWEB)

    Wodewotzky, T.I.; Lima-Neto, J.F. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Pereira-Júnior, O.C.M. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Departamento de Cirurgia e Anestesiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Sudano, M.J.; Lima, S.A.F. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Bersano, P.R.O. [Departamento de Patologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Yoshioka, S.A. [Instituto de Química de São Carlos, Universidade de São Paulo, São Carlos, SP (Brazil); Landim-Alvarenga, F.C. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil)

    2012-09-21

    Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs) to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA) as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF) on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium.

  17. Oxidized low-density lipoprotein-induced apoptotic dendritic cells as a novel therapy for atherosclerosis.

    Science.gov (United States)

    Frodermann, Vanessa; van Puijvelde, Gijs H M; Wierts, Laura; Lagraauw, H Maxime; Foks, Amanda C; van Santbrink, Peter J; Bot, Ilze; Kuiper, Johan; de Jager, Saskia C A

    2015-03-01

    Modulation of immune responses may form a powerful approach to treat atherosclerosis. It was shown that clearance of apoptotic cells results in tolerance induction to cleared Ags by dendritic cells (DCs); however, this seems impaired in atherosclerosis because Ag-specific tolerance is lacking. This could result, in part, from decreased emigration of DCs from atherosclerotic lesions because of the high-cholesterol environment. Nonetheless, local induction of anti-inflammatory responses by apoptotic cell clearance seems to dampen atherosclerosis, because inhibition of apoptotic cell clearance worsens atherosclerosis. In this study, we assessed whether i.v. administration of oxLDL-induced apoptotic DCs (apop(ox)-DCs) and, as a control, unpulsed apoptotic DCs could modulate atherosclerosis by inducing tolerance. Adoptive transfer of apop(ox)-DCs into low-density lipoprotein receptor knockout mice either before or during feeding of a Western-type diet resulted in increased numbers of CD103(+) tolerogenic splenic DCs, with a concomitant increase in regulatory T cells. Interestingly, both types of apoptotic DCs induced an immediate 40% decrease in Ly-6C(hi) monocyte numbers and a 50% decrease in circulating CCL2 levels, but only apop(ox)-DC treatment resulted in long-term effects on monocytes and CCL2 levels. Although initial lesion development was reduced by 40% in both treatment groups, only apop(ox)-DC treatment prevented lesion progression by 28%. Moreover, progressed lesions of apop(ox)-DC-treated mice showed a robust 45% increase in collagen content, indicating an enhanced stability of lesions. Our findings clearly show that apoptotic DC treatment significantly decreases lesion development, but only apop(ox)-DCs can positively modulate lesion progression and stability. These findings may translate into a safe treatment for patients with established cardiovascular diseases using patient-derived apop(ox)-DCs. Copyright © 2015 by The American Association of

  18. High density lipoprotein (HDL promotes glucose uptake in adipocytes and glycogen synthesis in muscle cells.

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    Qichun Zhang

    Full Text Available BACKGROUND: High density lipoprotein (HDL was reported to decrease plasma glucose and promote insulin secretion in type 2 diabetes patients. This investigation was designed to determine the effects and mechanisms of HDL on glucose uptake in adipocytes and glycogen synthesis in muscle cells. METHODS AND RESULTS: Actions of HDL on glucose uptake and GLUT4 translocation were assessed with 1-[(3H]-2-deoxyglucose and plasma membrane lawn, respectively, in 3T3-L1 adipocytes. Glycogen analysis was performed with amyloglucosidase and glucose oxidase-peroxidase methods in normal and palmitate-treated L6 cells. Small interfering RNA was used to observe role of scavenger receptor type I (SR-BI in glucose uptake of HDL. Corresponding signaling molecules were detected by immunoblotting. HDL stimulated glucose uptake in a time- and concentration-dependent manner in 3T3-L1 adipocytes. GLUT4 translocation was significantly increased by HDL. Glycogen deposition got enhanced in L6 muscle cells paralleling with elevated glycogen synthase kinase3 (GSK3 phosphorylation. Meanwhile, increased phosphorylations of Akt-Ser473 and AMP activated protein kinase (AMPK α were detected in 3T3-L1 adipocytes. Glucose uptake and Akt-Ser473 activation but not AMPK-α were diminished in SR-BI knock-down 3T3-L1 cells. CONCLUSIONS: HDL stimulates glucose uptake in 3T3-L1 adipocytes through enhancing GLUT4 translocation by mechanisms involving PI3K/Akt via SR-BI and AMPK signaling pathways, and increases glycogen deposition in L6 muscle cells through promoting GSK3 phosphorylation.

  19. Engineering biomechanically functional neocartilage derived from expanded articular chondrocytes through the manipulation of cell-seeding density and dexamethasone concentration.

    Science.gov (United States)

    Huang, Brian J; Huey, Daniel J; Hu, Jerry C; Athanasiou, Kyriacos A

    2017-08-01

    Recent work has established methods to engineer self-assembled, scaffold-free neocartilage from an expanded articular chondrocyte (AC) cell source. In continuing such work, the objective of the present study was to investigate the effects of cell-seeding density and dexamethasone concentration on these neocartilage constructs. Neocartilage discs (5 mm diameter) were formed by self-assembling passaged leporine articular chondrocytes into non-adherent agarose moulds. The cell-seeding densities (2, 3, 4, 5 and 6 million cells/construct) and dexamethasone concentrations (10 and 100 nm) in the culture medium were varied in a full-factorial study. After 4 weeks, the neocartilage constructs were assessed for morphological, biochemical and biomechanical properties. The cell-seeding density profoundly affected neocartilage properties. The two dexamethasone concentrations explored did not induce overall significant differences. Constructs formed using lower cell-seeding densities possessed much higher biochemical and biomechanical properties than constructs seeded with higher cell densities. Notably, the 2 million cells/construct group formed hyaline-like neocartilage with a collagen wet weight (WW) content of ~7% and a Young's modulus of ~4 MPa, representing the high end of values achieved in self-assembled neocartilage. Excitingly, the mechanical properties of these constructs were on a par with that of native cartilage tissues tested under similar conditions. Through optimization of cell-seeding density, this study shows for the first time the use of expanded ACs to form homogeneous self-assembled neocartilage with exceptionally high tensile strength. With such functional properties, these engineered neocartilage constructs provide a promising alternative for treating articular lesions. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  20. Impact of Pancreatic Rat Islet Density on Cell Survival during Hypoxia

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    A. Rodriguez-Brotons

    2016-01-01

    Full Text Available In bioartificial pancreases (BP, the number of islets needed to restore normoglycaemia in the diabetic patient is critical. However, the confinement of a high quantity of islets in a limited space may impact islet survival, particularly in regard to the low oxygen partial pressure (PO2 in such environments. The aim of the present study was to evaluate the impact of islet number in a confined space under hypoxia on cell survival. Rat islets were seeded at three different concentrations (150, 300, and 600 Islet Equivalents (IEQ/cm2 and cultured in normal atmospheric pressure (160 mmHg as well as hypoxic conditions (15 mmHg for 24 hours. Cell viability, function, hypoxia-induced changes in gene expression, and cytokine secretion were then assessed. Notably, hypoxia appeared to induce a decrease in viability and increasing islet density exacerbated the observed increase in cellular apoptosis as well as the loss of function. These changes were also associated with an increase in inflammatory gene transcription. Taken together, these data indicate that when a high number of islets are confined to a small space under hypoxia, cell viability and function are significantly impacted. Thus, in order to improve islet survival in this environment during transplantation, oxygenation is of critical importance.

  1. Whole-cell analysis of low-density lipoprotein uptake by macrophages using STEM tomography.

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    Jean-Pierre Baudoin

    Full Text Available Nanoparticles of heavy materials such as gold can be used as markers in quantitative electron microscopic studies of protein distributions in cells with nanometer spatial resolution. Studying nanoparticles within the context of cells is also relevant for nanotoxicological research. Here, we report a method to quantify the locations and the number of nanoparticles, and of clusters of nanoparticles inside whole eukaryotic cells in three dimensions using scanning transmission electron microscopy (STEM tomography. Whole-mount fixed cellular samples were prepared, avoiding sectioning or slicing. The level of membrane staining was kept much lower than is common practice in transmission electron microscopy (TEM, such that the nanoparticles could be detected throughout the entire cellular thickness. Tilt-series were recorded with a limited tilt-range of 80° thereby preventing excessive beam broadening occurring at higher tilt angles. The 3D locations of the nanoparticles were nevertheless determined with high precision using computation. The obtained information differed from that obtained with conventional TEM tomography data since the nanoparticles were highlighted while only faint contrast was obtained on the cellular material. Similar as in fluorescence microscopy, a particular set of labels can be studied. This method was applied to study the fate of sequentially up-taken low-density lipoprotein (LDL conjugated to gold nanoparticles in macrophages. Analysis of a 3D reconstruction revealed that newly up-taken LDL-gold was delivered to lysosomes containing previously up-taken LDL-gold thereby forming onion-like clusters.

  2. Automated computation of arbor densities: a step toward identifying neuronal cell types

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    Uygar eSümbül

    2014-11-01

    Full Text Available The shape and position of a neuron convey information regarding its molecular and functional identity. The identification of cell types from structure, a classic method, relies on the time-consuming step of arbor tracing. However, as genetic tools and imaging methods make data-driven approaches to neuronal circuit analysis feasible, the need for automated processing increases. Here, we first establish that mouse retinal ganglion cell types can be as precise about distributing their arbor volumes across the inner plexiform layer as they are about distributing the skeletons of the arbors. Then, we describe an automated approach to computing the spatial distribution of the dendritic arbors, or arbor density, with respect to a global depth coordinate based on this observation. Our method involves three-dimensional reconstruction of neuronal arbors by a supervised machine learning algorithm, post-processing of the enhanced stacks to remove somata and isolate the neuron of interest, and registration of neurons to each other using automatically detected arbors of the starburst amacrine interneurons as fiducial markers. In principle, this method could be generalizable to other structures of the CNS, provided that they allow sparse labeling of the cells and contain a reliable axis of spatial reference.

  3. Positive selection of Wharton's jelly-derived CD105(+) cells by MACS technique and their subsequent cultivation under suspension culture condition: A simple, versatile culturing method to enhance the multipotentiality of mesenchymal stem cells.

    Science.gov (United States)

    Amiri, Fatemeh; Halabian, Raheleh; Dehgan Harati, Mozhgan; Bahadori, Marzie; Mehdipour, Ahmad; Mohammadi Roushandeh, Amaneh; Habibi Roudkenar, Mehryar

    2015-05-01

    Wharton's jelly (WJ), an appropriate source of mesenchymal stem cells (MSCs), has been shown to have a wide array of therapeutic applications. However, the WJ-derived MSCs are very heterogeneous and have limited expression of pluripotency markers. Hence, improvement of their culture condition would promote the efficiency of WJ-MSCs. This study aims to employ a simple method of cultivation to obtain WJ-MSCs which express more pluripotency markers. CD105(+) cells were separated by magnetic-associated (activated) cell sorting from umbilical cord mucous tissue. CD105(+) cells were added to Methocult medium diluted in α-minimum essential medium (α-MEM) and seeded in poly(2-hydroxyethyl methacrylate) (poly-HEMA)-coated plates for suspension culture preparation. Differentiation capacity of isolated cells was evaluated in the presence of differentiation-inducing media. The expression of pluripotency markers such as Oct3/4, Nanog, and Sox2 was also analyzed by RT-PCR and western blot techniques. Moreover, immunocytochemistry was performed to detect alpha-smooth muscle actin (antigene) (α-SMA) protein. WJ-MSCs grew homogeneously and formed colonies when cultured under suspension culture conditions (Non-adhesive WJ-MSCs). They maintained their growth ability in both adherent and suspension cultures for several passages. Non-adhesive WJ-MSCs expressed Oct3/4, Nanog, and Sox2 both at transcriptional and translational levels in comparison to those cultured in conventional adherent cultures. They also expressed α-SMA protein. In this study, we isolated WJ-MSCs using a slightly modified culture condition. Our simple non-genetic method resulted in a homogeneous population of WJ-MSCs, which highly expressed pluripotency markers. In the future, more multipotent WJ-MSCs can be harnessed as a non-embryonic source of MSCs in MSC-based cell therapy.

  4. The Impact of Cathode Material and Shape on Current Density in an Aluminum Electrolysis Cell

    Science.gov (United States)

    Song, Yang; Peng, Jianping; Di, Yuezhong; Wang, Yaowu; Li, Baokuan; Feng, Naixiang

    2016-02-01

    A finite element model was developed to determine the impact of cathode material and shape on current density in an aluminum electrolysis cell. For the cathode material, results show that increased electrical resistivity leads to a higher cathode voltage drop; however, the horizontal current is reduced in the metal. The horizontal current magnitude for six different cathode materials in decreasing order is graphitized, semi-graphitized, full graphitic, 50% anthracite (50% artificial graphite), 70% anthracite (30% artificial graphite), 100% anthracite. The modified cathode shapes with an inclined cathode surface, higher collector bar and cylindrical protrusions are intended to improve horizontal current and flow resistance. Compared to a traditional cathode, modified collector bar sizes of 70 mm × 230 mm and 80 mm × 270 mm can reduce horizontal current density component Jx by 10% and 19%, respectively, due to better conductivity of the steel. The horizontal current in the metal decreases with increase of cathode inclination. The peak value of Jx can be approximately reduced by 20% for a 2° change in inclination. Cylindrical protrusions lead to local horizontal current increase on their tops, but the average current is less affected and the molten metal is effectively slowed down.

  5. Isolation of Peroxisomes from Rat Liver and Cultured Hepatoma Cells by Density Gradient Centrifugation.

    Science.gov (United States)

    Manner, Andreas; Islinger, Markus

    2017-01-01

    Subcellular fractionation is still a valuable technique to unravel organelle-specific proteomes, validate the location of uncharacterized proteins, or to functionally analyze import and metabolism in individual subcellular compartments. In this respect, density gradient centrifugation still represents a very classic, indispensable technique to isolate and analyze peroxisomes. Here, we present two independent protocols for the purification of peroxisomes from either liver tissue or the HepG2 hepatoma cell line. While the former permits the isolation of highly pure peroxisomes suitable for, e.g., subcellular proteomics experiments, the latter protocol yields peroxisomal fractions from considerably less purity but allows to easily modify metabolic conditions in the culture medium or to genetically manipulate the peroxisomal compartment. In this respect, both purification methods represent alternative tools to be applied in experiments investigating peroxisome physiology.

  6. Leptin deficiency-induced obesity affects the density of mast cells in abdominal fat depots and lymph nodes in mice

    Directory of Open Access Journals (Sweden)

    Altintas Mehmet M

    2012-02-01

    Full Text Available Abstract Background Mast cells are implicated in the pathogenesis of obesity and insulin resistance. Here, we explored the effects of leptin deficiency-induced obesity on the density of mast cells in metabolic (abdominal fat depots, skeletal muscle, and liver and lymphatic (abdominal lymph nodes, spleen, and thymus organs. Fourteen-week-old male leptin-deficient ob/ob mice and their controls fed a standard chow were studied. Tissue sections were stained with toluidine blue to determine the density of mast cells. CD117/c-kit protein expression analysis was also carried out. Furthermore, mast cells containing immunoreactive tumor necrosis factor-α (TNF-α, a proinflammatory cytokine involved in obesity-linked insulin resistance, were identified by immunostaining. Results ob/ob mice demonstrated adiposity and insulin resistance. In abdominal fat depots, mast cells were distributed differentially. While most prevalent in subcutaneous fat in controls, mast cells were most abundant in epididymal fat in ob/ob mice. Leptin deficiency-induced obesity was accompanied by a 20-fold increase in the density of mast cells in epididymal fat, but a 13-fold decrease in subcutaneous fat. This finding was confirmed by CD117/c-kit protein expression analysis. Furthermore, we found that a subset of mast cells in epididymal and subcutaneous fat were immunoreactive for TNF-α. The proportion of mast cells immunoreactive for TNF-α was higher in epididymal than in subcutaneous fat in both ob/ob and control mice. Mast cells were also distributed differentially in retroperitoneal, mesenteric, and inguinal lymph nodes. In both ob/ob mice and lean controls, mast cells were more prevalent in retroperitoneal than in mesenteric and inguinal lymph nodes. Leptin deficiency-induced obesity was accompanied by increased mast cell density in all lymph node stations examined. No significant difference in the density of mast cells in skeletal muscle, liver, spleen, and thymus was

  7. Possible Prognostic and Therapeutic Significance of c-Kit Expression, Mast Cell Count and Microvessel Density in Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Ilaria Marech

    2014-07-01

    Full Text Available Renal cell carcinoma (RCC is the most frequent renal tumor and its incidence is increasing worldwide. Tumor angiogenesis is known to play a crucial role in the etiopathogenesis of RCC and over the last few years an even deeper knowledge of its contribution in metastatic RCC development has led to the development of numerous molecular targeting agents (such as sunitinib, sorafenib, pazopanib, axitinib, tivozanib, and dovitinib. The above agents are principally directed against vascular endothelial growth factor receptor (VEGFR members and also against c-Kit receptor (c-KitR. The role of c-kitR inhibition on clear cell RCC (ccRCC, the main RCC subtype, is less well established. Whether c-kitR activation through its ligand, stem cell factor (SCF contributes significantly to the effects of tyrosine kinase inhibitors (TKIs treatment remains to be established. It is important to underscore that the c-KitR is expressed on mast cells (MCs and cancer cells. After an examination of the c-KitR/SCF pathway, we review here the principal studies that have evaluated c-Kit expression in RCC. Moreover, we summarize some investigations that have observed the distribution of MCs in primary renal cancer and in adjacent normal tissue with appropriate histological immunohistochemical techniques. We also focus on few studies that have evaluated the correlation between RCC proliferation, MC count and microvessel density (MVD, as hallmarks of tumor angiogenesis. Thus, the aim of this review of the literature is to clarify if c-KitR expression, MC count and MVD could have prognostic significance and the possible predictive therapeutic implications in RCC.

  8. [Dendrobium officinale stereoscopic cultivation method].

    Science.gov (United States)

    Si, Jin-Ping; Dong, Hong-Xiu; Liao, Xin-Yan; Zhu, Yu-Qiu; Li, Hui

    2014-12-01

    The study is aimed to make the most of available space of Dendrobium officinale cultivation facility, reveal the yield and functional components variation of stereoscopic cultivated D. officinale, and improve quality, yield and efficiency. The agronomic traits and yield variation of stereoscopic cultivated D. officinale were studied by operating field experiment. The content of polysaccharide and extractum were determined by using phenol-sulfuric acid method and 2010 edition of "Chinese Pharmacopoeia" Appendix X A. The results showed that the land utilization of stereoscopic cultivated D. officinale increased 2.74 times, the stems, leaves and their total fresh or dry weight in unit area of stereoscopic cultivated D. officinale were all heavier than those of the ground cultivated ones. There was no significant difference in polysaccharide content between stereoscopic cultivation and ground cultivation. But the extractum content and total content of polysaccharide and extractum were significantly higher than those of the ground cultivated ones. In additional, the polysaccharide content and total content of polysaccharide and extractum from the top two levels of stereoscopic culture matrix were significantly higher than that of the ones from the other levels and ground cultivation. Steroscopic cultivation can effectively improves the utilization of space and yield, while the total content of polysaccharides and extractum were significantly higher than that of the ground cultivated ones. The significant difference in Dendrobium polysaccharides among the plants from different height of stereo- scopic culture matrix may be associated with light factor.

  9. High-Density and Very-Low-Density Lipoprotein Have Opposing Roles in Regulating Tumor-Initiating Cells and Sensitivity to Radiation in Inflammatory Breast Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Wolfe, Adam R. [Department of Radiation Oncology, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Morgan Welch Inflammatory Breast Cancer Research Program and Clinic, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Atkinson, Rachel L. [Morgan Welch Inflammatory Breast Cancer Research Program and Clinic, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Reddy, Jay P. [Department of Radiation Oncology, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Debeb, Bisrat G.; Larson, Richard; Li, Li [Department of Radiation Oncology, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Department of Clinical Cancer Prevention, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Masuda, Hiroko; Brewer, Takae [Department of Clinical Cancer Prevention, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Atkinson, Bradley J. [Department of Clinical Pharmacy Services, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Brewster, Abeena [Morgan Welch Inflammatory Breast Cancer Research Program and Clinic, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Ueno, Naoto T. [Department of Clinical Cancer Prevention, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Woodward, Wendy A., E-mail: wwoodward@mdanderson.org [Department of Radiation Oncology, University of Texas MD Anderson Cancer Center, Houston, Texas (United States); Department of Clinical Cancer Prevention, University of Texas MD Anderson Cancer Center, Houston, Texas (United States)

    2015-04-01

    Purpose: We previously demonstrated that cholesterol-lowering agents regulate radiation sensitivity of inflammatory breast cancer (IBC) cell lines in vitro and are associated with less radiation resistance among IBC patients who undergo postmastectomy radiation. We hypothesized that decreasing IBC cellular cholesterol induced by treatment with lipoproteins would increase radiation sensitivity. Here, we examined the impact of specific transporters of cholesterol (ie lipoproteins) on the responses of IBC cells to self-renewal and to radiation in vitro and on clinical outcomes in IBC patients. Methods and Materials: Two patient-derived IBC cell lines, SUM 149 and KPL4, were incubated with low-density lipoproteins (LDL), very-low-density lipoproteins (VLDL), or high-density lipoproteins (HDL) for 24 hours prior to irradiation (0-6 Gy) and mammosphere formation assay. Cholesterol panels were examined in a cohort of patients with primary IBC diagnosed between 1995 and 2011 at MD Anderson Cancer Center. Lipoprotein levels were then correlated to patient outcome, using the log rank statistical model, and examined in multivariate analysis using Cox regression. Results: VLDL increased and HDL decreased mammosphere formation compared to untreated SUM 149 and KPL4 cells. Survival curves showed enhancement of survival in both of the IBC cell lines when pretreated with VLDL and, conversely, radiation sensitization in all cell lines when pretreated with HDL. In IBC patients, higher VLDL values (>30 mg/dL) predicted a lower 5-year overall survival rate than normal values (hazard ratio [HR] = 1.9 [95% confidence interval [CI]: 1.05-3.45], P=.035). Lower-than-normal patient HDL values (<60 mg/dL) predicted a lower 5-year overall survival rate than values higher than 60 mg/dL (HR = 3.21 [95% CI: 1.25-8.27], P=.015). Conclusions: This study discovered a relationship among the plasma levels of lipoproteins, overall patient response, and radiation resistance in IBC patients

  10. Increased COX-2 expression in epithelial and stromal cells of high mammographic density tissues and in a xenograft model of mammographic density.

    Science.gov (United States)

    Chew, G L; Huo, C W; Huang, D; Hill, P; Cawson, J; Frazer, H; Hopper, J L; Haviv, I; Henderson, M A; Britt, K; Thompson, E W

    2015-08-01

    Mammographic density (MD) adjusted for age and body mass index is one of the strongest known risk factors for breast cancer. Given the high attributable risk of MD for breast cancer, chemoprevention with a safe and available agent that reduces MD and breast cancer risk would be beneficial. Cox-2 has been implicated in MD-related breast cancer risk, and was increased in stromal cells in high MD tissues in one study. Our study assessed differential Cox-2 expression in epithelial and stromal cells in paired samples of high and low MD human breast tissue, and in a validated xenograft biochamber model of MD. We also examined the effects of endocrine treatment upon Cox-2 expression in high and low MD tissues in the MD xenograft model. Paired high and low MD human breast tissue samples were immunostained for Cox-2, then assessed for differential expression and staining intensity in epithelial and stromal cells. High and low MD human breast tissues were separately maintained in biochambers in mice treated with Tamoxifen, oestrogen or placebo implants, then assessed for percentage Cox-2 staining in epithelial and stromal cells. Percentage Cox-2 staining was greater for both epithelial (p = 0.01) and stromal cells (p tissues. In high MD biochamber tissues, percentage Cox-2 staining was greater in stromal cells of oestrogen-treated versus placebo-treated tissues (p = 0.05).

  11. Cell density dependence of Microcystis aeruginosa responses to copper algaecide concentrations: Implications for microcystin-LR release.

    Science.gov (United States)

    Kinley, Ciera M; Iwinski, Kyla J; Hendrikse, Maas; Geer, Tyler D; Rodgers, John H

    2017-11-01

    Along with mechanistic models, predictions of exposure-response relationships for copper are often derived from laboratory toxicity experiments with standardized experimental exposures and conditions. For predictions of copper toxicity to algae, cell density is a critical factor often overlooked. For pulse exposures of copper-based algaecides in aquatic systems, cell density can significantly influence copper sorbed by the algal population, and consequent responses. A cyanobacterium, Microcystis aeruginosa, was exposed to a copper-based algaecide over a range of cell densities to model the density-dependence of exposures, and effects on microcystin-LR (MC-LR) release. Copper exposure concentrations were arrayed to result in a gradient of MC-LR release, and masses of copper sorbed to algal populations were measured following exposures. While copper exposure concentrations eliciting comparable MC-LR release ranged an order of magnitude (24-h EC50s 0.03-0.3mg Cu/L) among cell densities of 10 6 through 10 7 cells/mL, copper doses (mg Cu/mg algae) were similar (24-h EC50s 0.005-0.006mg Cu/mg algae). Comparisons of MC-LR release as a function of copper exposure concentrations and doses provided a metric of the density dependence of algal responses in the context of copper-based algaecide applications. Combined with estimates of other site-specific factors (e.g. water characteristics) and fate processes (e.g. dilution and dispersion, sorption to organic matter and sediments), measuring exposure-response relationships for specific cell densities can refine predictions for in situ exposures and algal responses. These measurements can in turn decrease the likelihood of amending unnecessary copper concentrations to aquatic systems, and minimize risks for non-target aquatic organisms. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Goat and sheep ovarian tissue cryopreservation: Effects on the morphology and development of primordial follicles and density of stromal cell.

    Science.gov (United States)

    Faustino, L R; Santos, R R; Silva, C M G; Pinto, L C; Celestino, J J H; Campello, C C; Figueiredo, J R; Rodrigues, A P R

    2010-10-01

    The effect of exposure to cryoprotectant and cryopreservation of goat and sheep ovarian cortical fragments on the morphology of primordial follicles, stromal cell density and follicular development was performed. Goat and sheep ovarian fragments were exposed to 1.0 or 1.5M ethylene glycol (EG) for 5, 10 or 20min, followed or not by conventional cryopreservation. Follicular morphology and stromal cell density were evaluated by means of classical histological analysis. In addition, ovarian fragments were cultured for 1 or 7 days after cryopreservation to evaluate follicular development. Both exposure to cryoprotectant and cryopreservation of goat and sheep ovarian tissue did affect the morphology of primordial follicles and stromal cell density, except when goat ovarian tissue was exposed to EG for 5min. Although exposure time did not influence follicular morphology in both species, increase in the exposure time from 5 to 20min did reduce goat stromal cell density. Increase in EG concentration from 1.0 to 1.5M did result in the decrease of the percentage of goat morphologically normal primordial follicles evaluated after exposure only. In vitro culture of frozen-thawed goat and sheep ovarian tissue showed that exposure to 1.0M, for 10min, before freezing of goat and sheep ovarian tissue does not impair follicular developmental capacity. In addition, stromal cell density may play a role in follicular survival and development after cryopreservation of ovarian tissue. Copyright © 2010 Elsevier B.V. All rights reserved.

  13. Cultivating strategic thinking skills.

    Science.gov (United States)

    Shirey, Maria R

    2012-06-01

    This department highlights change management strategies that may be successful in strategically planning and executing organizational change initiatives. With the goal of presenting practical approaches helpful to nurse leaders advancing organizational change, content includes evidence-based projects, tools, and resources that mobilize and sustain organizational change initiatives. In this article, the author presents an overview of strategic leadership and offers approaches for cultivating strategic thinking skills.

  14. Cultivating an entrepreneurial mindset.

    Science.gov (United States)

    Matheson, Sandra A

    2013-01-01

    Now as never before, familiar challenges require bold, novel approaches. Registered dietitians will benefit by cultivating an entrepreneurial mindset that involves being comfortable with uncertainty, learning to take calculated risks, and daring to just try it. An entrepreneur is someone who takes risks to create something new, usually in business. But the entrepreneurial mindset is available to anyone prepared to rely only on their own abilities for their economic security and expect no opportunity without first creating value for others.

  15. Cultivating Leaders of Indiana

    OpenAIRE

    yaryyeva, Annagul; Sdunzik, Jennifer

    2016-01-01

    "Cultivating Leaders of Indiana" was developed to establish connections between the Purdue student body and the Frankfort community. By engaging high school students in workshops that focused on local, national, and global identities, the goal of the project was to encourage students to appreciate their individuality and to motivate them to translate their skills into a global perspective.Moreover, workshops centering on themes such as culture, citizenship, media, and education were designed ...

  16. Establishing elements of a synthetic biology platform for Vaccinia virus production: BioBrick™ design, serum-free virus production and microcarrier-based cultivation of CV-1 cells

    Directory of Open Access Journals (Sweden)

    Shuchang Liu

    2017-02-01

    Full Text Available Vaccinia virus (VACV is an established vector for vaccination and is beginning to prove effective as an oncolytic agent. Industrial production of VACV stands to benefit in future from advances made by synthetic biology in genome engineering and standardisation. The CV-1 cell line can be used for VACV propagation and has been used extensively with the CRISPR/Cas9 system for making precise edits of the VACV genome. Here we take first steps toward establishing a scalable synthetic biology platform for VACV production with CV-1 cells featuring standardised biological tools and serum free cell cultivation. We propose a new BioBrick™ plasmid backbone format for inserting transgenes into VACV. We then test the performance of CV-1 cells in propagation of a conventional recombinant Lister strain VACV, VACVL-15 RFP, in a serum-free process. CV-1 cells grown in 5% foetal bovine serum (FBS Dulbecco’s Modified Eagle Medium (DMEM were adapted to growth in OptiPRO and VP-SFM brands of serum-free media. Specific growth rates of 0.047 h−1 and 0.044 h−1 were observed for cells adapted to OptiPRO and VP-SFM respectively, compared to 0.035 h−1 in 5% FBS DMEM. Cells adapted to OptiPRO and to 5% FBS DMEM achieved recovery ratios of over 96%, an indication of their robustness to cryopreservation. Cells adapted to VP-SFM showed a recovery ratio of 82%. Virus productivity in static culture, measured as plaque forming units (PFU per propagator cell, was 75 PFU/cell for cells in 5% FBS DMEM. VP-SFM and OptiPRO adaptation increased VACV production to 150 PFU/cell and 350 PFU/cell respectively. Boosted PFU/cell from OptiPRO-adapted cells persisted when 5% FBS DMEM or OptiPRO medium was observed during the infection step and when titre was measured using cells adapted to 5% FBS DMEM or OptiPRO medium. Finally, OptiPRO-adapted CV-1 cells were successfully cultivated using Cytodex-1 microcarriers to inform future scale up studies.

  17. Establishing elements of a synthetic biology platform for Vaccinia virus production: BioBrick™ design, serum-free virus production and microcarrier-based cultivation of CV-1 cells.

    Science.gov (United States)

    Liu, Shuchang; Ruban, Ludmila; Wang, Yaohe; Zhou, Yuhong; Nesbeth, Darren N

    2017-02-01

    Vaccinia virus (VACV) is an established vector for vaccination and is beginning to prove effective as an oncolytic agent. Industrial production of VACV stands to benefit in future from advances made by synthetic biology in genome engineering and standardisation. The CV-1 cell line can be used for VACV propagation and has been used extensively with the CRISPR/Cas9 system for making precise edits of the VACV genome. Here we take first steps toward establishing a scalable synthetic biology platform for VACV production with CV-1 cells featuring standardised biological tools and serum free cell cultivation. We propose a new BioBrick™ plasmid backbone format for inserting transgenes into VACV. We then test the performance of CV-1 cells in propagation of a conventional recombinant Lister strain VACV, VACVL-15 RFP, in a serum-free process. CV-1 cells grown in 5% foetal bovine serum (FBS) Dulbecco's Modified Eagle Medium (DMEM) were adapted to growth in OptiPRO and VP-SFM brands of serum-free media. Specific growth rates of 0.047 h-1 and 0.044 h-1 were observed for cells adapted to OptiPRO and VP-SFM respectively, compared to 0.035 h-1 in 5% FBS DMEM. Cells adapted to OptiPRO and to 5% FBS DMEM achieved recovery ratios of over 96%, an indication of their robustness to cryopreservation. Cells adapted to VP-SFM showed a recovery ratio of 82%. Virus productivity in static culture, measured as plaque forming units (PFU) per propagator cell, was 75 PFU/cell for cells in 5% FBS DMEM. VP-SFM and OptiPRO adaptation increased VACV production to 150 PFU/cell and 350 PFU/cell respectively. Boosted PFU/cell from OptiPRO-adapted cells persisted when 5% FBS DMEM or OptiPRO medium was observed during the infection step and when titre was measured using cells adapted to 5% FBS DMEM or OptiPRO medium. Finally, OptiPRO-adapted CV-1 cells were successfully cultivated using Cytodex-1 microcarriers to inform future scale up studies.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2015-10-30

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

  19. Paralytic shellfish toxin concentration and cell density changes in Pyrodinium bahamense -Noctiluca scintillans feeding experiments.

    Science.gov (United States)

    Azanza, Rhodora V; Cruz, Lourdes J; Cariño, Flerida A; Blanco, Alelea G; Butardo, Vito M

    2010-05-01

    For the first time the potential of Noctiluca scintillans, a non-toxic mixotrophic dinoflagellate, in bioconverting and/or excreting saxitoxin has been illustrated, thus contributing to the limited knowledge on the aspects of toxin pathways in the food chain/web and predator-prey preferences. Noctiluca growth rate increased with higher Pyrodinium concentration but the ratio of Noctiluca to Pyrodinium should at least be 1:250 cells per mL. Noctiluca fed with Pyrodinium alone was found to decrease in number suggesting that the nutrients from this prey were insufficient. This was confirmed by the improved cell density of Noctiluca upon addition of 0.01% casitone to the Pyrodinium-fed Noctiluca. The alternative prey (Gymnodinium sanguineum) slowed down the grazing impact of Noctiluca on Pyrodinium. Noctiluca depleted Gymnodinium earlier than Pyrodinium showing preference over a prey with less saxitoxin. After the feeding experiments, total saxitoxin levels decreased to 72% in the Noctiluca-Pyrodinium setup whereas no saxitoxin was detected in the Noctiluca culture fed with Pyrodinium and G. sanguineum. It is possible that Gymnodinium can provide some nutrients needed to make Noctiluca more efficient in bioconverting saxitoxin. Copyright 2009 Elsevier Ltd. All rights reserved.

  20. Optimized LTE cell planning for multiple user density subareas using meta-heuristic algorithms

    KAUST Repository

    Ghazzai, Hakim

    2014-09-01

    Base station deployment in cellular networks is one of the most fundamental problems in network design. This paper proposes a novel method for the cell planning problem for the fourth generation 4G-LTE cellular networks using meta heuristic algorithms. In this approach, we aim to satisfy both coverage and cell capacity constraints simultaneously by formulating a practical optimization problem. We start by performing a typical coverage and capacity dimensioning to identify the initial required number of base stations. Afterwards, we implement a Particle Swarm Optimization algorithm or a recently-proposed Grey Wolf Optimizer to find the optimal base station locations that satisfy both problem constraints in the area of interest which can be divided into several subareas with different user densities. Subsequently, an iterative approach is executed to eliminate eventual redundant base stations. We have also performed Monte Carlo simulations to study the performance of the proposed scheme and computed the average number of users in outage. Results show that our proposed approach respects in all cases the desired network quality of services even for large-scale dimension problems.

  1. Optimized LTE Cell Planning with Varying Spatial and Temporal User Densities

    KAUST Repository

    Ghazzai, Hakim

    2015-03-09

    Base station deployment in cellular networks is one of the fundamental problems in network design. This paper proposes a novel method for the cell planning problem for the fourth generation (4G) cellular networks using meta-heuristic algorithms. In this approach, we aim to satisfy both cell coverage and capacity constraints simultaneously by formulating an optimization problem that captures practical planning aspects. The starting point of the planning process is defined through a dimensioning exercise that captures both coverage and capacity constraints. Afterwards, we implement a meta-heuristic algorithm based on swarm intelligence (e.g., particle swarm optimization or the recently-proposed grey wolf optimizer) to find suboptimal base station locations that satisfy both problem constraints in the area of interest which can be divided into several subareas with different spatial user densities. Subsequently, an iterative approach is executed to eliminate eventual redundant base stations. We also perform Monte Carlo simulations to study the performance of the proposed scheme and compute the average number of users in outage. Next, the problems of green planning with regards to temporal traffic variation and planning with location constraints due to tight limits on electromagnetic radiations are addressed, using the proposed method. Finally, in our simulation results, we apply our proposed approach for different scenarios with different subareas and user distributions and show that the desired network quality of service targets are always reached even for large-scale problems.

  2. High-density lipoprotein-mediated transcellular cholesterol transport in mouse aortic endothelial cells.

    Science.gov (United States)

    Miao, LiXia; Okoro, Emmanuel U; Cao, ZhiJan; Yang, Hong; Motley-Johnson, Evangeline; Guo, Zhongmao

    2015-09-18

    Accumulation of unesterified cholesterol-rich lipid vesicles in the subendothelial space contributes to atherogenesis. Transport of cholesterol from the subendothelial intima back to the circulating blood inhibits atherosclerosis development; however, the mechanism for this process has not been fully defined. Using cultured mouse aortic endothelial cells (MAECs), we observed that unesterified cholesterol can be transported across the endothelial cell monolayer from the basolateral to the apical compartment. Administration of high-density lipoprotein (HDL) or apolipoprotein AI (apoAI) to the apical compartment enhanced transendothelial cholesterol transport in a concentration-dependent manner. Knockdown of ATP-binding cassette transporter G1 (ABCG1) or scavenger receptor class B type I (SR-B1), or inhibition of SR-B1 diminished HDL-induced transendothelial cholesterol transport; while knockdown of ABCA1 reduced apoAI-mediated cholesterol transport. HDL enhanced phosphorylation of phosphatidylinositol 3-kinase (PI3K) and Akt in MAECs. However, inhibition of PI3K or Akt did not reduce HDL-induced transendothelial cholesterol transport. These results suggest that HDL enhances transendothelial cholesterol transport by activation of a mechanism involving ABCA1, ABCG1 and SR-B1 but not involving PI3K and Akt. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. High cell-density production of poly(3-hydroxybutyrate) in a membrane bioreactor.

    Science.gov (United States)

    Haas, Cornelia; El-Najjar, Tarek; Virgolini, Nikolaus; Smerilli, Marina; Neureiter, Markus

    2017-07-25

    Agro-industrial residues with a low carbon content, such as whey, stillage or wastewater from plant oil mills are abundant and cheap. However, they cannot be used directly in highly productive industrial poly(3-hydroxybutyrate) (P3HB) production, as the classical fed-batch fermentation strategy requires highly concentrated feed streams. This problem has been circumvented in this report by retaining the cells during the fermentation in the bioreactor using an external microfiltration module. Synthetic medium containing a glucose concentration of 50g/L was continuously fed to Cupriavidus necator, which converted the sugar to P3HB. With this setup we were able to achieve high productivities (3.10g P3HB/(Lh)) and reach high cell densities (148g/L) containing 76% P3HB, and obtained good yields (0.33g P3HB/g added glucose). The added sugar from the feed was instantly consumed by the bacteria, resulting in a negligible loss of sugar to the permeate. This approach creates the possibility of polyhydroxyalkanoate production from a range of cheap and easily available substrates, for which only waste water treatment or biogas production has been cost-competitive until now. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Increased extracellular matrix density decreases MCF10A breast cell acinus formation in 3D culture conditions.

    Science.gov (United States)

    Lance, Amanda; Yang, Chih-Chao; Swamydas, Muthulekha; Dean, Delphine; Deitch, Sandy; Burg, Karen J L; Dréau, Didier

    2016-01-01

    The extracellular matrix (ECM) contributes to the generation and dynamic of normal breast tissue, in particular to the generation of polarized acinar and ductal structures. In vitro 3D culture conditions, including variations in the composition of the ECM, have been shown to directly influence the formation and organization of acinus-like and duct-like structures. Furthermore, the density of the ECM appears to also play a role in the normal mammary tissue and tumour formation. Here we show that the density of the ECM directly influences the number, organization and function of breast acini. Briefly, non-malignant human breast MCF10A cells were incubated in increasing densities of a Matrigel®-collagen I matrix. Elastic moduli near and distant to the acinus structures were measured by atomic force microscopy, and the number of acinus structures was determined. Immunochemistry was used to investigate the expression levels of E-cadherin, laminin, matrix metalloproteinase-14 and ß-casein in MCF10A cells. The modulus of the ECM was significantly increased near the acinus structures and the number of acinus structures decreased with the increase in Matrigel-collagen I density. As evaluated by the expression of laminin, the organization of the acinus structures present was altered as the density of the ECM increased. Increases in both E-cadherin and MMP14 expression by MCF10A cells as ECM density increased were also observed. In contrast, MCF10A cells expressed lower ß-casein levels as the ECM density increased. Taken together, these observations highlight the key role of ECM density in modulating the number, organization and function of breast acini. Copyright © 2013 John Wiley & Sons, Ltd.

  5. Culture density influence on the photosynthetic efficiency of microalgae growing under different spectral compositions of light.

    Science.gov (United States)

    Kula, M; Kalaji, H M; Skoczowski, A

    2017-02-01

    A density in algal suspension causes a significant change in the intensity and spectral composition of light reaching individual cells. Measurements of chlorophyll fluorescence allow us to observe any general changes in the bioenergetic status of photosynthesis. The aim of the study was to determine the effect of cultivation density on the PSII photochemical efficiency of three species of algae (Chlorella vulgaris, Botryococcus braunii and Chlorella emersonii), each with a different rate of growth - high, medium and low - respectively. The cell density of algae in suspension differentiated through the cultivation time (2, 4, and 8days) and the spectral composition of light. The results showed that the density of cultivation led to change in the photosynthetic apparatus of algae. The differences described between each day of cultivation (2, 4, and 8) in the kinetics of chlorophyll a fluorescence intensity in cells of the algal strains under study probably resulted from the different phases of growth of these cultures. In addition the results showed the beneficial effect of far red light on the photosynthetic apparatus and the growth of biomass in investigated algal strains. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Nicotine-induced expression of low-density lipoprotein receptor in oral epithelial cells.

    Directory of Open Access Journals (Sweden)

    Satoshi Ito

    Full Text Available BACKGROUND: Nicotine use is one of the most important risk factors for the development of cardiovascular and periodontal diseases. Numerous reports have suggested the possible contribution of disturbed lipid metabolism for the development of both disease groups. Despite these observations, little is known about the relationship between tobacco smoking and the development of these diseases. Our previous microarray data revealed that nicotine induced low-density lipoprotein receptor (LDLR expression in oral epithelial cells (OECs. The aim of the present study was to confirm nicotine-mediated LDLR induction and to elucidate the signaling mechanisms leading to the augmented expression of LDLR in OECs. METHODS AND RESULTS: LDLR and nicotinic acetylcholine receptor (nAChR subunit expression was detected by real-time PCR. The production of LDLR was demonstrated by immunofluorescence staining. nAChR-mediated LDLR induction was examined by pre-incubation of the cells with its specific inhibitor, α-bungarotoxin (α-BTX. The functional importance of transcription factor specific protein 1 (Sp1 was examined by luciferase assay, mithramycin pre-incubation or by small interfering RNA (siRNA transfection. The specific binding of Sp1 to R3 region of LDLR 5'-untranslated region was demonstrated with electrophoretic mobility shift assay (EMSA and streptavidin-agarose precipitation assay followed by western blotting. The results confirmed that nicotine induced LDLR expression at the transcriptional level. Nicotine was sensed by nAChR and the signal was transduced by Sp1 which bound to the R3 region of LDLR gene. Augmented production of LDLR in the gingival epithelial cells was further demonstrated by immunofluorescence staining using the gingival tissues obtained from the smoking patients. CONCLUSIONS: Taken together, the results suggested that nicotine might contribute to the development of both cardiovascular and periodontal diseases by inducing the LDLR in

  7. Plasmodium falciparum exhibits markers of regulated cell death at high population density in vitro.

    Science.gov (United States)

    Engelbrecht, Dewaldt; Coetzer, Thérèsa Louise

    2016-12-01

    The asexual erythrocytic cycle of the protozoan parasite Plasmodium falciparum is responsible for the pathogenesis of malaria and causes the overwhelming majority of malaria deaths. Rapidly increasing parasitaemia during this 48hour cycle threatens the survival of the human host and the parasite prior to transmission of the slow-maturing sexual stages to the mosquito host. The parasite may utilise regulated cell death (RCD) to control the burden of infection on the host and thus aid its own survival and transmission. The occurrence of RCD in P. falciparum remains a controversial topic. We provide strong evidence for the occurrence of an apoptosis-like phenotype of RCD in P. falciparum under conditions of high parasite density. P. falciparum was maintained in vitro and stressed by allowing growth to an unrestricted peak parasitaemia. Cell death markers, including morphological changes, DNA fragmentation, mitochondrial polarisation and phosphatidylserine externalisation were used to characterise parasite death at the time of peak parasitaemia and 24h later. At peak parasitaemia, mitochondrial depolarisation was observed, together with phosphatidylserine externalisation in both parasitised- and neighbouring non-infected erythrocytes. DNA fragmentation coincided with a decline in parasitaemia. Fewer merozoites were observed in mature schizonts at peak parasitaemia. Growth recovery to near-peak parasitaemia was noted within two intraerythrocytic cycles. The combination and chronological order of the biochemical markers of cell death suggest the occurrence of an apoptosis-like phenotype. The identification of a RCD pathway in P. falciparum may provide novel drug targets, particularly if the pathway differs from the host machinery. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  8. 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.

  9. Nitrate concentration-shift cultivation to enhance protein content of heterotrophic microalga Chlorella vulgaris: Over-compensation strategy.

    Science.gov (United States)

    Xie, Tonghui; Xia, Yun; Zeng, Yu; Li, Xingrui; Zhang, Yongkui

    2017-06-01

    Protein production from microalgae requires both high cell density during cultivation and high protein content in cells. Heterotrophic microalgae can achieve high cell density, and yet are confronted with the problem of low protein content. Based on over-compensation strategy, a new concentration-shift method was proposed to cultivate heterotrophic Chlorella vulgaris, aiming to increase protein content. With a prior starvation period, microalgae utilized more nitrate and accumulated more proteins compared to one-stage cultivation. Considering the convenience of operation, nitrate-added culture was adopted for producing heterotrophic microalgae, rather than sterile centrifugal culture. Operating parameters including nitrate concentration in N-deficient medium, N-starved time and nitrate concentration in N-rich medium were optimized, which were 0.18gl(-1), 38h and 2.45gl(-1), respectively. Under the optimized conditions, protein content in heterotrophic Chlorella reached 44.3%. Furthermore, the heterotrophic microalga was suggested to be a potential single-cell protein source according to the amino acid composition. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Prefrontal cortical parvalbumin and somatostatin expression and cell density increase during adolescence and are modified by BDNF and sex.

    Science.gov (United States)

    Du, X; Serena, K; Hwang, W; Grech, A M; Wu, Y W C; Schroeder, A; Hill, R A

    2018-02-03

    Brain-derived neurotrophic factor (BDNF) is known to play a critical role early in the development of cortical GABAergic interneurons. Recently our laboratory and others have shown protracted development of specific subpopulations of GABAergic interneurons extending into adolescence. BDNF expression also changes significantly across adolescent development. However the role of BDNF in regulating GABAergic changes across adolescence remains unclear. Here, we performed a week-by-week analysis of the protein expression and cell density of three major GABAergic interneurons, parvalbumin (PV), somatostatin (SST) and calretinin (Cal) in the medial prefrontal cortex from prepubescence (week 3) to adulthood (week 12). In order to assess how BDNF and sex might influence the adolescent trajectory of GABAergic interneurons we compared WT as well as BDNF heterozygous (+/-) male and female mice. In both males and females PV expression increases during adolescent development in the mPFC. Compared to wild-types, PV expression was reduced in male but not female BDNF+/- mice throughout adolescent development. This reduction in protein expression corresponded with reduced cell density, specifically within the infralimbic prefrontal cortex. SST expression increased in early adolescent WT females and this upregulation was delayed in BDNF+/-. SST cell density also increased in early adolescent mPFC of WT female mice, with BDNF+/- again showing a reduced pattern of expression. Cal protein expression was also sex-dependently altered across adolescence with WT males showing a steady decline but that of BDNF+/- remaining unaltered. Reduced cell density in on the other hand was observed particularly in male BDNF+/- mice. In females, Cal protein expression and cell density remained largely stable. Our results show that PV, SST and calretinin interneurons are indeed still developing into early adolescence in the mPFC and that BDNF plays a critical, sex-specific role in mediating expression and

  11. Stomatal and pavement cell density linked to leaf internal CO2 concentration.

    Science.gov (United States)

    Santrůček, Jiří; Vráblová, Martina; Simková, Marie; Hronková, Marie; Drtinová, Martina; Květoň, Jiří; Vrábl, Daniel; Kubásek, Jiří; Macková, Jana; Wiesnerová, Dana; Neuwithová, Jitka; Schreiber, Lukas

    2014-08-01

    Stomatal density (SD) generally decreases with rising atmospheric CO2 concentration, Ca. However, SD is also affected by light, air humidity and drought, all under systemic signalling from older leaves. This makes our understanding of how Ca controls SD incomplete. This study tested the hypotheses that SD is affected by the internal CO2 concentration of the leaf, Ci, rather than Ca, and that cotyledons, as the first plant assimilation organs, lack the systemic signal. Sunflower (Helianthus annuus), beech (Fagus sylvatica), arabidopsis (Arabidopsis thaliana) and garden cress (Lepidium sativum) were grown under contrasting environmental conditions that affected Ci while Ca was kept constant. The SD, pavement cell density (PCD) and stomatal index (SI) responses to Ci in cotyledons and the first leaves of garden cress were compared. (13)C abundance (δ(13)C) in leaf dry matter was used to estimate the effective Ci during leaf development. The SD was estimated from leaf imprints. SD correlated negatively with Ci in leaves of all four species and under three different treatments (irradiance, abscisic acid and osmotic stress). PCD in arabidopsis and garden cress responded similarly, so that SI was largely unaffected. However, SD and PCD of cotyledons were insensitive to Ci, indicating an essential role for systemic signalling. It is proposed that Ci or a Ci-linked factor plays an important role in modulating SD and PCD during epidermis development and leaf expansion. The absence of a Ci-SD relationship in the cotyledons of garden cress indicates the key role of lower-insertion CO2 assimilation organs in signal perception and its long-distance transport. © The Author 2014. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  12. High power density microbial fuel cell with flexible 3D graphene-nickel foam as anode.

    Science.gov (United States)

    Wang, Hanyu; Wang, Gongming; Ling, Yichuan; Qian, Fang; Song, Yang; Lu, Xihong; Chen, Shaowei; Tong, Yexiang; Li, Yat

    2013-11-07

    The structure and electrical conductivity of anode play a significant role in the power generation of microbial fuel cells (MFCs). In this study, we developed a three-dimensional (3D) reduced graphene oxide-nickel (denoted as rGO-Ni) foam as an anode for MFC through controlled deposition of rGO sheets onto the nickel foam substrate. The loading amount of rGO sheets and electrode surface area can be controlled by the number of rGO loading cycles. 3D rGO-Ni foam anode provides not only a large accessible surface area for microbial colonization and electron mediators, but also a uniform macro-porous scaffold for effective mass diffusion of the culture medium. Significantly, at a steady state of the power generation, the MFC device with flexible rGO-Ni electrodes produced an optimal volumetric power density of 661 W m(-3) calculated based on the volume of anode material, or 27 W m(-3) based on the volume of the anode chamber. These values are substantially higher than that of plain nickel foam, and other conventional carbon based electrodes (e.g., carbon cloth, carbon felt, and carbon paper) measured in the same conditions. To our knowledge, this is the highest volumetric power density reported for mL-scale MFC device with a pure strain of Shewanella oneidensis MR-1. We also demonstrated that the MFC device can be operated effectively in a batch-mode at least for a week. These new 3D rGO-Ni electrodes show great promise for improving the power generation of MFC devices.

  13. A mathematical model of the maximum power density attainable in an alkaline hydrogen/oxygen fuel cell

    Science.gov (United States)

    Kimble, Michael C.; White, Ralph E.

    1991-01-01

    A mathematical model of a hydrogen/oxygen alkaline fuel cell is presented that can be used to predict the polarization behavior under various power loads. The major limitations to achieving high power densities are indicated and methods to increase the maximum attainable power density are suggested. The alkaline fuel cell model describes the phenomena occurring in the solid, liquid, and gaseous phases of the anode, separator, and cathode regions based on porous electrode theory applied to three phases. Fundamental equations of chemical engineering that describe conservation of mass and charge, species transport, and kinetic phenomena are used to develop the model by treating all phases as a homogeneous continuum.

  14. High cell density cultivation of Pseudomonas oleovorans : Growth and production of poly (3-hydroxyalkanoates) in two-liquid phase batch and fed-batch systems

    NARCIS (Netherlands)

    Preusting, Hans; VANHOUTEN, R; HOEFS, A; VANLANGENBERGHE, EK; FAVREBULLE, O; Witholt, Bernard

    1993-01-01

    Pseudomonas oleovorans is able to accumulate poly(3-hydroxyalkanoates) (PHAs) under conditions of excess n-alkanes, which serve as sole energy and carbon source, and limitation of an essential nutrient such as ammonium. In this study we aimed at an efficient production of these PHAs by growing P.

  15. Allelopathy as a potential strategy to improve microalgae cultivation

    National Research Council Canada - National Science Library

    Bacellar Mendes, Leonardo Brantes; Vermelho, Alane Beatriz

    2013-01-01

    One of the main obstacles for continuous productivity in microalgae cultivation is the presence of biological contaminants capable of eliminating large numbers of cells in a matter of days or even hours...

  16. Mycobacterial Cultures Contain Cell Size and Density Specific Sub-populations of Cells with Significant Differential Susceptibility to Antibiotics, Oxidative and Nitrite Stress

    Science.gov (United States)

    Vijay, Srinivasan; Nair, Rashmi Ravindran; Sharan, Deepti; Jakkala, Kishor; Mukkayyan, Nagaraja; Swaminath, Sharmada; Pradhan, Atul; Joshi, Niranjan V.; Ajitkumar, Parthasarathi

    2017-01-01

    The present study shows the existence of two specific sub-populations of Mycobacterium smegmatis and Mycobacterium tuberculosis cells differing in size and density, in the mid-log phase (MLP) cultures, with significant differential susceptibility to antibiotic, oxidative, and nitrite stress. One of these sub-populations (~10% of the total population), contained short-sized cells (SCs) generated through highly-deviated asymmetric cell division (ACD) of normal/long-sized mother cells and symmetric cell divisions (SCD) of short-sized mother cells. The other sub-population (~90% of the total population) contained normal/long-sized cells (NCs). The SCs were acid-fast stainable and heat-susceptible, and contained high density of membrane vesicles (MVs, known to be lipid-rich) on their surface, while the NCs possessed negligible density of MVs on the surface, as revealed by scanning and transmission electron microscopy. Percoll density gradient fractionation of MLP cultures showed the SCs-enriched fraction (SCF) at lower density (probably indicating lipid-richness) and the NCs-enriched fraction (NCF) at higher density of percoll fractions. While live cell imaging showed that the SCs and the NCs could grow and divide to form colony on agarose pads, the SCF, and NCF cells could independently regenerate MLP populations in liquid and solid media, indicating their full genomic content and population regeneration potential. CFU based assays showed the SCF cells to be significantly more susceptible than NCF cells to a range of concentrations of rifampicin and isoniazid (antibiotic stress), H2O2 (oxidative stress),and acidified NaNO2 (nitrite stress). Live cell imaging showed significantly higher susceptibility of the SCs of SC-NC sister daughter cell pairs, formed from highly-deviated ACD of normal/long-sized mother cells, to rifampicin and H2O2, as compared to the sister daughter NCs, irrespective of their comparable growth rates. The SC-SC sister daughter cell pairs, formed

  17. Functional evaluation of corneal endothelium by combined measurement of corneal volume alteration and cell density after phacoemulsification.

    Science.gov (United States)

    Suzuki, Hisaharu; Oki, Kotaro; Takahashi, Keizo; Shiwa, Toshihiko; Takahashi, Hiroshi

    2007-12-01

    To assess a new method of functional evaluation of corneal endothelium using combined measurement of corneal volume alteration and cell density (CD) after phacoemulsification. Nippon Medical School Hospital, Tokyo, Japan. Phacoemulsification was performed in 28 cases. The corneal volume within 3.0 mm and 10.0 mm circles of the cornea was measured using Scheimpflug imaging (Pentacam, Oculus, Inc.) and the cell density, using specular microscopy (Noncon Robo, Konan Medical). The volume stress index (VSI), a parameter for measuring functional evaluation, was calculated by dividing the volume increase within the 3.0 mm circle (preoperative corneal volume - corneal volume at time of measurement) by the central cell density value. The resulting value is indicative of the amount of postoperative increase in corneal volume with which each corneal endothelial cell should be able to cope. Three months after surgery, the 3.0 mm corneal volume returned to preoperative values, while the 10.0 mm corneal volume remained significantly higher than preoperatively. The cell density values did not significantly change throughout the study. The VSI showed a significant decrease by 7 days after surgery followed by a gradual decrease until the end of the study. At 3 months, the VSI value was significantly smaller than at 1 day. The corneal volume increase after surgery may be indicative of the degree of the endothelial damage in the area; thus, the VSI may be useful in the functional assessment of the corneal endothelium.

  18. Cell-density-dependent regulation of expression and glycosylation of dopachrome tautomerase/tyrosinase-related protein-2.

    Science.gov (United States)

    Hornyak, T J; Hayes, D J; Ziff, E B

    2000-07-01

    The expression of the dopachrome tautomerase gene (Dct) and its protein product, tyrosinase-related protein-2, was studied in the cultured, phorbol-ester-dependent murine melanocyte cell line melan-a. Increased cell density was found to stimulate Dct expression both in cells stably transfected with a Dct promoter-lacZ construct and endogenously in nontransfected cells. Increased Dct expression under these conditions corresponds to increased tyrosinase-related protein-2 production. Tyrosinase-related protein-2 was found to exist in two distinct glycoforms with different endoglycosidase sensitivities. Density-dependent expression of tyrosinase-related protein-2 was independent of time of cell growth, cell proliferation, and soluble factors, implying that cell-cell contact is the important determinant governing increased Dct expression under these conditions. Tyrp1 gene expression and tyrosinase-related protein-1 production were also induced under similar conditions. The results show that cell-cell contact between melanocytes induces a coordinated response at both transcriptional and nontranscriptional levels that induces production of the tyrosinase-related proteins that have a significant role in melanization.

  19. Low track height standard-cells enable high-placement density and low-BEOL cost (Conference Presentation)

    Science.gov (United States)

    Debacker, Peter; Matti, Luca; Sherazi, Syed M. Y.; Baert, Rogier; Gerousis, Vassilios; Nauts, Claire; Raghavan, Praveen; Ryckaert, Julien; Kim, Ryoung-Han; Verkest, Diederik

    2017-04-01

    Making standards cells smaller by lowering the cell height from 7.5 tracks to 6 tracks for the same set of ground rules is an efficient way to reduce area for high density digital IP blocks without increasing wafer cost. Denser cells however also imply a higher pin density and possible more routing congestion because of that. In Place and Route phase, this limits the cell density (a.k.a. utilization) that can be reached without design rule violations. This study shows that 6-track cells (192nm high) and smart routing results in up to 60% lower area than 7.5-track cells in N5 technology. Standard cells have been created for 7.5T and 6T cells in N5 technology (poly pitch 42nm, metal pitch 32nm). The cells use a first horizontal routing layer (Mint) and vertical M1 for 1D intra-cell routing as much as possible. Place and route was performed on an opencores LDPC decoder. Various cell architectures and place and route optimizations are used to scale down the cell area and improve density. Most are not process optimizations, but optimized cell architectures and routing methods: • Open M1: M1 is removed as much as possible. This allows the router to use M1 for inter-cell routing in dense areas. • Routing in Mint: With open M1 the router can also use Mint to extend pins to access nearby free M1 tracks in congested areas. • Outbound rail: The 7.5T cells have inbound VDD/VSS rails in Mint for easy supply tapping. Moving the Mint rail outbound and shared between cells is required to enable lower track height cells. • Vertical Power distribution network (PDN): in 6T cells too many horizontal tracks would be consumed by the wide M2 rail. Mint is used instead combined with a vertical PDN in M1. • Self-Aligned Gate Contact allows to contact the gate on top of active fins. Any Mint track then can contact a gate, reducing cell area considerably. • Partially landing Mint Via trench: In 6T cells, a continuous Via trench underneath the Mint rail is used. This via partially

  20. Lipid Droplets Characterization in Adipocyte Differentiated 3T3-L1 Cells: Size and Optical Density Distribution

    Science.gov (United States)

    Rizzatti, V.; Boschi, F.; Pedrotti, M.; Zoico, E.; Sbarbati, A.; Zamboni, M.

    2013-01-01

    The 3T3-L1 cell line, derived from 3T3 cells, is widely used in biological research on adipose tissue. 3T3-L1 cells have a fibroblast-like morphology, but, under appropriate conditions, they differentiate into an adipocyte-like phenotype. During the differentiation process, 3T3-L1 cells increase the synthesis of triglycerides and acquire the behavior of adipose cells. In particular, triglycerides accumulate in lipid droplets (LDs) embedded in the cytoplasm. The number and the size distribution of the LDs is often correlated with obesity and many other pathologies linked with fat accumulation. The integrated optical density (IOD) of the LDs is related with the amount of triglycerides in the droplets. The aim of this study is the attempt to characterize the size distribution and the IOD of the LDs in 3T3-L1 differentiated cells. The cells were differentiated into adipocytes for 5 days with a standard procedure, stained with Oil Red O and observed with an optical microscope. The diameter, area, optical density of the LDs were measured. We found an asymmetry of the kernel density distribution of the maximum Feret’s diameter of the LDs with a tail due to very large LDs. More information regarding the birth of the LDs could help in finding the best mathematical model in order to analyze fat accumulation in adipocytes. PMID:24085273

  1. Intratumoral CD3 and CD8 T-cell Densities Associated with Relapse-Free Survival in HCC.

    Science.gov (United States)

    Gabrielson, Andrew; Wu, Yunan; Wang, Hongkun; Jiang, Jiji; Kallakury, Bhaskar; Gatalica, Zoran; Reddy, Sandeep; Kleiner, David; Fishbein, Thomas; Johnson, Lynt; Island, Eddie; Satoskar, Rohit; Banovac, Filip; Jha, Reena; Kachhela, Jaydeep; Feng, Perry; Zhang, Tiger; Tesfaye, Anteneh; Prins, Petra; Loffredo, Christopher; Marshall, John; Weiner, Louis; Atkins, Michael; He, Aiwu Ruth

    2016-05-01

    Immune cells that infiltrate a tumor may be a prognostic factor for patients who have had surgically resected hepatocellular carcinoma (HCC). The density of intratumoral total (CD3(+)) and cytotoxic (CD8(+)) T lymphocytes was measured in the tumor interior and in the invasive margin of 65 stage I to IV HCC tissue specimens from a single cohort. Immune cell density in the interior and margin was converted to a binary score (0, low; 1, high), which was correlated with tumor recurrence and relapse-free survival (RFS). In addition, the expression of programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1) was correlated with the density of CD3(+) and CD8(+) cells and clinical outcome. High densities of both CD3(+) and CD8(+) T cells in both the interior and margin, along with corresponding Immunoscores, were significantly associated with a low rate of recurrence (P = 0.007) and a prolonged RFS (P = 0.002). In multivariate logistic regression models adjusted for vascular invasion and cellular differentiation, both CD3(+) and CD8(+) cell densities predicted recurrence, with odds ratios of 5.8 [95% confidence interval (CI), 1.6-21.8] for CD3(+) and 3.9 (95% CI, 1.1-14.1) for CD8(+) Positive PD-L1 staining was correlated with high CD3 and CD8 density (P = 0.024 and 0.005, respectively) and predicted a lower rate of recurrence (P = 0.034), as well as prolonged RFS (P = 0.029). Immunoscore and PD-L1 expression, therefore, are useful prognostic markers in patients with HCC who have undergone primary tumor resection. Cancer Immunol Res; 4(5); 419-30. ©2016 AACR. ©2016 American Association for Cancer Research.

  2. Argyrophil cell density in the oxyntic mucosa is higher in female than in male morbidly obese patients

    Directory of Open Access Journals (Sweden)

    F.A.N. Maksud

    2013-05-01

    Full Text Available Obesity is a multifactorial disorder often associated with many important diseases such as diabetes, hypertension and other metabolic syndrome conditions. Argyrophil cells represent almost the total population of endocrine cells of the human gastric mucosa and some reports have described changes of specific types of these cells in patients with obesity and metabolic syndrome. The present study was designed to evaluate the global population of argyrophil cells of the gastric mucosa of morbidly obese and dyspeptic non-obese patients. Gastric biopsies of antropyloric and oxyntic mucosa were obtained from 50 morbidly obese patients (BMI >40 and 50 non-obese patients (17 dyspeptic overweight and 33 lean individuals and processed for histology and Grimelius staining for argyrophil cell demonstration. Argyrophil cell density in the oxyntic mucosa of morbidly obese patients was higher in female (238.68 ± 83.71 cells/mm2 than in male patients (179.31 ± 85.96 cells/mm2 and also higher in female (214.20 ± 50.38 cells/mm2 than in male (141.90 ± 61.22 cells/mm2 morbidly obese patients with metabolic syndrome (P = 0.01 and P = 0.02, respectively. In antropyloric mucosa, the main difference in argyrophil cell density was observed between female morbidly obese patients with (167.00 ± 69.30 cells/mm2 and without (234.00 ± 69.54 cells/mm2 metabolic syndrome (P = 0.001. In conclusion, the present results show that the number of gastric argyrophil cells could be under gender influence in patients with morbid obesity. In addition, gastric argyrophil cells seem to behave differently among female morbidly obese patients with and without metabolic syndrome.

  3. Argyrophil cell density in the oxyntic mucosa is higher in female than in male morbidly obese patients

    Directory of Open Access Journals (Sweden)

    F.A.N. Maksud

    2013-04-01

    Full Text Available Obesity is a multifactorial disorder often associated with many important diseases such as diabetes, hypertension and other metabolic syndrome conditions. Argyrophil cells represent almost the total population of endocrine cells of the human gastric mucosa and some reports have described changes of specific types of these cells in patients with obesity and metabolic syndrome. The present study was designed to evaluate the global population of argyrophil cells of the gastric mucosa of morbidly obese and dyspeptic non-obese patients. Gastric biopsies of antropyloric and oxyntic mucosa were obtained from 50 morbidly obese patients (BMI >40 and 50 non-obese patients (17 dyspeptic overweight and 33 lean individuals and processed for histology and Grimelius staining for argyrophil cell demonstration. Argyrophil cell density in the oxyntic mucosa of morbidly obese patients was higher in female (238.68 ± 83.71 cells/mm2 than in male patients (179.31 ± 85.96 cells/mm2 and also higher in female (214.20 ± 50.38 cells/mm2 than in male (141.90 ± 61.22 cells/mm2 morbidly obese patients with metabolic syndrome (P = 0.01 and P = 0.02, respectively. In antropyloric mucosa, the main difference in argyrophil cell density was observed between female morbidly obese patients with (167.00 ± 69.30 cells/mm2 and without (234.00 ± 69.54 cells/mm2 metabolic syndrome (P = 0.001. In conclusion, the present results show that the number of gastric argyrophil cells could be under gender influence in patients with morbid obesity. In addition, gastric argyrophil cells seem to behave differently among female morbidly obese patients with and without metabolic syndrome.

  4. Argyrophil cell density in the oxyntic mucosa is higher in female than in male morbidly obese patients

    Energy Technology Data Exchange (ETDEWEB)

    Maksud, F.A.N. [Laboratório de Patologia Digestiva e Neuroendócrina, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil); Faculdade de Medicina, Universidade Federal de Ouro Preto, Ouro Preto, MG (Brazil); Kakehasi, A.M. [Laboratório de Patologia Digestiva e Neuroendócrina, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil); Barbosa, A.J.A. [Laboratório de Patologia Digestiva e Neuroendócrina, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil); Instituto Alfa de Gastroenterologia, Belo Horizonte, MG (Brazil)

    2013-04-05

    Obesity is a multifactorial disorder often associated with many important diseases such as diabetes, hypertension and other metabolic syndrome conditions. Argyrophil cells represent almost the total population of endocrine cells of the human gastric mucosa and some reports have described changes of specific types of these cells in patients with obesity and metabolic syndrome. The present study was designed to evaluate the global population of argyrophil cells of the gastric mucosa of morbidly obese and dyspeptic non-obese patients. Gastric biopsies of antropyloric and oxyntic mucosa were obtained from 50 morbidly obese patients (BMI >40) and 50 non-obese patients (17 dyspeptic overweight and 33 lean individuals) and processed for histology and Grimelius staining for argyrophil cell demonstration. Argyrophil cell density in the oxyntic mucosa of morbidly obese patients was higher in female (238.68 ± 83.71 cells/mm{sup 2}) than in male patients (179.31 ± 85.96 cells/mm{sup 2}) and also higher in female (214.20 ± 50.38 cells/mm{sup 2}) than in male (141.90 ± 61.22 cells/mm{sup 2}) morbidly obese patients with metabolic syndrome (P = 0.01 and P = 0.02, respectively). In antropyloric mucosa, the main difference in argyrophil cell density was observed between female morbidly obese patients with (167.00 ± 69.30 cells/mm{sup 2}) and without (234.00 ± 69.54 cells/mm{sup 2}) metabolic syndrome (P = 0.001). In conclusion, the present results show that the number of gastric argyrophil cells could be under gender influence in patients with morbid obesity. In addition, gastric argyrophil cells seem to behave differently among female morbidly obese patients with and without metabolic syndrome.

  5. Enhanced xylose fermentation by engineered yeast expressing NADH oxidase through high cell density inoculums.

    Science.gov (United States)

    Zhang, Guo-Chang; Turner, Timothy L; Jin, Yong-Su

    2017-03-01

    Accumulation of reduced byproducts such as glycerol and xylitol during xylose fermentation by engineered Saccharomyces cerevisiae hampers the economic production of biofuels and chemicals from cellulosic hydrolysates. In particular, engineered S. cerevisiae expressing NADPH-linked xylose reductase (XR) and NAD + -linked xylitol dehydrogenase (XDH) produces substantial amounts of the reduced byproducts under anaerobic conditions due to the cofactor difference of XR and XDH. While the additional expression of a water-forming NADH oxidase (NoxE) from Lactococcus lactis in engineered S. cerevisiae with the XR/XDH pathway led to reduced glycerol and xylitol production and increased ethanol yields from xylose, volumetric ethanol productivities by the engineered yeast decreased because of growth defects from the overexpression of noxE. In this study, we introduced noxE into an engineered yeast strain (SR8) exhibiting near-optimal xylose fermentation capacity. To overcome the growth defect caused by the overexpression of noxE, we used a high cell density inoculum for xylose fermentation by the SR8 expressing noxE. The resulting strain, SR8N, not only showed a higher ethanol yield and lower byproduct yields, but also exhibited a high ethanol productivity during xylose fermentation. As noxE overexpression elicits a negligible growth defect on glucose conditions, the beneficial effects of noxE overexpression were substantial when a mixture of glucose and xylose was used. Consumption of glucose led to rapid cell growth and therefore enhanced the subsequent xylose fermentation. As a result, the SR8N strain produced more ethanol and fewer byproducts from a mixture of glucose and xylose than the parental SR8 strain without noxE overexpression. Our results suggest that the growth defects from noxE overexpression can be overcome in the case of fermenting lignocellulose-derived sugars such as glucose and xylose.

  6. Investigation of the Electron Acceleration by a High-Power Laser and a Density-Tapered Mixed-Gas Cell

    Science.gov (United States)

    Kim, Jinju; Phung, Vanessa L. J.; Kim, Minseok; Hur, Min-Sup; Suk, Hyyong

    2017-10-01

    Plasma-based accelerators can generate about 1000 times stronger acceleration field compared with RF-based conventional accelerators, which can be done by high power laser and plasma. There are many issues in this research and one of them is development of a good plasma source for higher electron beam energy. For this purpose, we are investigating a special type of plasma source, which is a density-tapered gas cell with a mixed-gas for easy injection. By this type of special gas cell, we expect higher electron beam energies with easy injection in the wakefield. In this poster, some experimental results for electron beam generation with the density-tapered mixed-gas cell are presented. In addition to the experimental results, CFD (Computational-Fluid-Dynamics) and PIC (Particle-In-Cell) simulation results are also presented for comparison studies.

  7. Expression of low density lipoprotein receptor-related protein 4 (Lrp4) gene in the mouse germ cells.

    Science.gov (United States)

    Yamaguchi, Yasuka L; Tanaka, Satomi S; Kasa, Miyuki; Yasuda, Kunio; Tam, Patrick P L; Matsui, Yasuhisa

    2006-08-01

    The low density lipoprotein receptor-related protein 4 gene (Lrp4) was identified by subtractive screening of cDNAs of the migratory primordial germ cells (PGCs) of E8.5-9.5 embryo and E3.5 blastocysts. Lrp4 is expressed in PGCs in the hindgut and the dorsal mesentery of E9.5 embryos, and in germ cells in the genital ridges of male and female E10.5-13.5 embryos. Lrp4 is also expressed in spermatogonia of the neonatal and adult testes and in the immature oocytes and follicular cells of the adult ovary. The absence of Lrp4 expression in the blastocyst, embryonic stem cells and embryonic germ cells suggests the Lrp4 is a molecular marker that distinguishes the germ cells from embryo-derived pluripotent stem cells.

  8. LOW-TEMPERATURE, ANODE-SUPPORTED HIGH POWER DENSITY SOLID OXIDE FUEL CELLS WITH NANOSTRUCTURED ELECTRODES

    Energy Technology Data Exchange (ETDEWEB)

    Professor Anil V. Virkar

    2003-05-23

    This report summarizes the work done during the entire project period, between October 1, 1999 and March 31, 2003, which includes a six-month no-cost extension. During the project, eight research papers have, either been, published, accepted for publication, or submitted for publication. In addition, several presentations have been made in technical meetings and workshops. The project also has provided support for four graduate students working towards advanced degrees. The principal technical objective of the project was to analyze the role of electrode microstructure on solid oxide fuel cell performance. Prior theoretical work conducted in our laboratory demonstrated that the particle size of composite electrodes has a profound effect on cell performance; the finer the particle size, the lower the activation polarization, the better the performance. The composite cathodes examined consisted of electronically conducting perovskites such as Sr-doped LaMnO{sub 3} (LSM) or Sr-doped LaCoO{sub 3} (LSC), which is also a mixed conductor, as the electrocatalyst, and yttria-stabilized zirconia (YSZ) or rare earth oxide doped CeO{sub 2} as the ionic conductor. The composite anodes examined were mixtures of Ni and YSZ. A procedure was developed for the synthesis of nanosize YSZ by molecular decomposition, in which unwanted species were removed by leaching, leaving behind nanosize YSZ. Anode-supported cells were made using the as-synthesized powders, or using commercially acquired powders. The electrolyte was usually a thin ({approx}10 microns), dense layer of YSZ, supported on a thick ({approx}1 mm), porous Ni + YSZ anode. The cathode was a porous mixture of electrocatalyst and an ionic conductor. Most of the cell testing was done at 800 C with hydrogen as fuel and air as the oxidant. Maximum power densities as high as 1.8 W/cm{sup 2} were demonstrated. Polarization behavior of the cells was theoretically analyzed. A limited amount of cell testing was done using liquid

  9. Secretory expression of functional barley limit dextrinase by Pichia pastoris using high cell-density fermentation

    DEFF Research Database (Denmark)

    Vester-Christensen, Malene Bech; Abou Hachem, Maher; Næsted, Henrik

    2010-01-01

    Heterologous production of large multidomain proteins from higher plants is often cumbersome. Barley limit dextrinase (LD), a 98 kDa multidomain starch and alpha-limit dextrin debranching enzyme, plays a major role in starch mobilization during seed germination and is possibly involved in starch ...... catalytic coefficient, k(cat,app)/K-m,K-app = 488 +/- 23 mL/(mg s) is 3.5-fold higher than for barley malt LD. Surface plasmon resonance analysis showed alpha-, beta-, and gamma-cyclodextrin binding to LD with K-d of 27.2, 0.70, and 34.7 mu M, respectively.......Heterologous production of large multidomain proteins from higher plants is often cumbersome. Barley limit dextrinase (LD), a 98 kDa multidomain starch and alpha-limit dextrin debranching enzyme, plays a major role in starch mobilization during seed germination and is possibly involved in starch...... biosynthesis by trimming of intermediate branched alpha-glucan structures. Highly active barley LD is obtained by secretory expression during high cell-density fermentation of Pichia pastoris. The LD encoding gene fragment without signal peptide was subcloned in-frame with the Saccharomyces cerevisiae alpha...

  10. Cryoprotective effects of low-density lipoproteins, trehalose and soybean lecithin on murine spermatogonial stem cells.

    Science.gov (United States)

    Wang, Peng; Li, Ying; Hu, Xiao-Chen; Cai, Xiao-Li; Hou, Li-Peng; Wang, Yan-Feng; Hu, Jian-Hong; Li, Qing-Wang; Suo, Li-Juan; Fan, Zhi-Guo; Zhang, Bo

    2014-05-01

    Spermatogonial stem cells (SSCs) have the ability to self-renew and offer a pathway for genetic engineering of the male germ line. Cryopreservation of SSCs has potential value for the treatment of male infertility, spermatogonial transplantation, and so on. In order to investigate the cryopreservation effects of different cryoprotectants on murine SSCs, 0.2 M of low-density lipoproteins (LDL), trehalose and soybean lecithin were added to the cryoprotective medium, respectively, and the murine SSCs were frozen at -80°C or -196°C. The results indicated that the optimal recovery rates of murine SSCs in the cryoprotective medium supplemented with LDL, trehalose and soybean lecithin were 92.53, 76.35 and 75.48% at -80°C, respectively. Compared with freezing at -196°C, the optimum temperature for improvement of recovery rates of frozen murine SSCs, cryopreservation in three different cryoprotectants at -80°C, were 17.11, 6.68 and 10.44% respectively. The recovery rates of murine SSCs in the cryoprotective medium supplemented with 0.2 M LDL were significantly higher than that of other cryoprotectants (P membranes. However, more research is needed to evaluate and understand the precise role of LDL during the freezing-thawing of SSCs.

  11. Shewanella oneidensis Hfq promotes exponential phase growth, stationary phase culture density, and cell survival

    Directory of Open Access Journals (Sweden)

    Brennan Christopher M

    2013-02-01

    Full Text Available Abstract Background Hfq is an RNA chaperone protein that has been broadly implicated in sRNA function in bacteria. Here we describe the construction and characterization of a null allele of the gene that encodes the RNA chaperone Hfq in Shewanella oneidensis strain MR-1, a dissimilatory metal reducing bacterium. Results Loss of hfq in S. oneidensis results in a variety of mutant phenotypes, all of which are fully complemented by addition of a plasmid-borne copy of the wild type hfq gene. Aerobic cultures of the hfq∆ mutant grow more slowly through exponential phase than wild type cultures, and hfq∆ cultures reach a terminal cell density in stationary phase that is ~2/3 of that observed in wild type cultures. We have observed a similar growth phenotype when the hfq∆ mutant is cultured under anaerobic conditions with fumarate as the terminal electron acceptor, and we have found that the hfq∆ mutant is defective in Cr(VI reduction. Finally, the hfq∆ mutant exhibits a striking loss of colony forming units in extended stationary phase and is highly sensitive to oxidative stress induced by H2O2 or methyl viologen (paraquat. Conclusions The hfq mutant in S. oneidensis exhibits pleiotropic phenotypes, including a defect in metal reduction. Our results also suggest that hfq mutant phenotypes in S. oneidensis may be at least partially due to increased sensitivity to oxidative stress.

  12. PORTSMOUTH ON-SITE DISPOSAL CELL HIGH DENSITY POLYETHYLENE GEOMEMBRANE LONGEVITY

    Energy Technology Data Exchange (ETDEWEB)

    Phifer, M.

    2012-01-31

    It is anticipated that high density polyethylene (HDPE) geomembranes will be utilized within the liner and closure cap of the proposed On-Site Disposal Cell (OSDC) at the Portsmouth Gaseous Diffusion Plant. The likely longevity (i.e. service life) of HDPE geomembranes in OSDC service is evaluated within the following sections of this report: (1) Section 2.0 provides an overview of HDPE geomembranes, (2) Section 3.0 outlines potential HDPE geomembranes degradation mechanisms, (3) Section 4.0 evaluates the applicability of HDPE geomembrane degradation mechanisms to the Portsmouth OSDC, (4) Section 5.0 provides a discussion of the current state of knowledge relative to the longevity (service life) of HDPE geomembranes, including the relation of this knowledge to the Portsmouth OSDC, and (5) Section 6.0 provides summary and conclusions relative to the anticipated service life of HDPE geomembranes in OSDC service. Based upon this evaluation it is anticipated that the service life of HDPE geomembranes in OSDC service would be significantly greater than the 200 year service life assumed for the OSDC closure cap and liner HDPE geomembranes. That is, a 200 year OSDC HDPE geomembrane service life is considered a conservative assumption.

  13. Systematic development and optimization of chemically defined medium supporting high cell density growth of Bacillus coagulans.

    Science.gov (United States)

    Chen, Yu; Dong, Fengqing; Wang, Yonghong

    2016-09-01

    With determined components and experimental reducibility, the chemically defined medium (CDM) and the minimal chemically defined medium (MCDM) are used in many metabolism and regulation studies. This research aimed to develop the chemically defined medium supporting high cell density growth of Bacillus coagulans, which is a promising producer of lactic acid and other bio-chemicals. In this study, a systematic methodology combining the experimental technique with flux balance analysis (FBA) was proposed to design and simplify a CDM. The single omission technique and single addition technique were employed to determine the essential and stimulatory compounds, before the optimization of their concentrations by the statistical method. In addition, to improve the growth rationally, in silico omission and addition were performed by FBA based on the construction of a medium-size metabolic model of B. coagulans 36D1. Thus, CDMs were developed to obtain considerable biomass production of at least five B. coagulans strains, in which two model strains B. coagulans 36D1 and ATCC 7050 were involved.

  14. Time dependent – density functional theory characterization of organic dyes for dye-sensitized solar cells

    KAUST Repository

    Hilal, Rifaat

    2017-06-19

    We aim at providing better insight into the parameters that govern the intramolecular charge transfer (ICT) and photo-injection processes in dyes for dye-sensitised solar cells (DSSC). Density functional theory (DFT) and time-dependent DFT (TD-DFT) calculations are utilized to study the geometry, electronic structure, electrostatic potential (ESP) and absorption spectrum, for a representative donor-π bridge-acceptor (D–π–A) dye for DSSC. The coplanar geometry of the dye (D1) facilitates strong conjugation and considerable delocalization originating the π CT interaction from donor to acceptor orbitals and the hyper-conjugative interactions involving Rydberg states. A model simulating the adsorption of the dye on the TiO surface is utilized to estimate binding energies. The effect of fluorine substituents in the π-spacer on the quantum efficiency of DSSCs was investigated. Gibb’s free energy values, redox potentials, excited state lifetime, non-linear optical properties (NLO) and driving forces for D1 and its fluorinated derivatives were computed.

  15. Bone mineral density, vitamin D, and nutritional status of children submitted to hematopoietic stem cell transplantation.

    Science.gov (United States)

    Campos, Denise Johnsson; Boguszewski, César Luiz; Funke, Vaneuza Araujo Moreira; Bonfim, Carmem Maria Sales; Kulak, Carolina Aguiar Moreira; Pasquini, Ricardo; Borba, Victória Zeghbi Cochenski

    2014-06-01

    The aim of the study was to evaluate the effect of allogeneic hematopoietic stem cell transplantation (HSCT) on bone mineral density (BMD), serum vitamin D levels, and nutritional status of 50 patients between ages 4 and 20 y. We conducted pre-HSCT and 6-mo post-HSCT evaluations. We measured BMD at the lumbar spine (LS) and total body (TB) by dual energy x-ray absorptiometry (DXA); body composition by bioimpedance analysis, and dietary intakes of calcium and vitamin D using the 24-h recall and semiquantitative food frequency questionnaire methods. We observed a significant reduction in BMD 6 mo post-HSCT. Nearly half (48%) of patients had reductions at the LS (average -9.6% ± 6.0%), and patients who developed graft-versus-host disease (GVHD) had the greatest reductions (-5.6% versus 1.2%, P nutritional status, and vitamin D levels. We suggest that early routine assessment be done to permit prevention and treatment. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Effect of orientation and density of hydroxide precursor films on performance of dye-sensitized ZnO solar cells

    National Research Council Canada - National Science Library

    KAJIHARA, Hisasuke; HAGIWARA, Manabu; FUJIHARA, Shinobu

    2016-01-01

    .... The increase in the film density resulted in the improvement of the open-circuit voltage and the fill factor of dye-sensitized solar cells using N719-loaded ZnO films as photoanodes. The light-to-electricity conversion efficiency of 3.49% was obtained in the DSSC using the present ZnO film.

  17. Secretory otitis. Histopathology and goblet-cell density in the Eustachian tube and middle ear in children.

    Science.gov (United States)

    Tos, M; Bak-Pedersen, K

    1976-05-01

    The histological changes in the middle ear of four children with incipient or mild chronic secretory otitis are described. They consist in vascular dilatation and proliferation, round-cell infiltration, epithelial metaplasia into pseudostratified, columnar, ciliated epithelium posteriorly in the middle ear, formation of abnormal mucous tubular glands, and an increase in goblet-cell density in the osseous tube and middle ear. The causes of these changes were presumably long-lasting tubal occlusion due to a permanent nasogastric tube and protracted catarrhal diseases. The density of goblet cells was determined in various parts of the Eustachian tube and middle ear. The findings were analysed statistically and compared with the density in prematures and newborns, normal children, and normal adults. In the tympanic orifice and in the osseous Eustachian tube the goblet-cell density was greatly increased, whereas the increase in the middle ear varied individually, but was in conformity with the other findings. These cases illustrate that the histopathological changes in the middle-ear mucosa in secretory otitis must be regarded not only from a qualitative, but certainly also from a quantitative point of view.

  18. Magnetic Resonance Imaging Detection of Tumor Cells by Targeting Low-Density Lipoprotein Receptors with Gd-Loaded Low-Density Lipoprotein Particles

    Directory of Open Access Journals (Sweden)

    Simonetta Geninatti Crich

    2007-12-01

    Full Text Available Gd-DO3A-diph and Gd-AAZTAC17 are lipophilic magnetic resonance imaging (MRI agents that display high affinity for low-density lipoprotein (LDL particles. However, on binding to LDL, Gd-DO3A-diph shows a decreased hydration that results in a lower enhancement of water proton relaxation rate. Conversely, GdAAZTAC17 displays a strong relaxation enhancement at the imaging fields. Each LDL particle can load up to 100 and 400 UNITS of Gd-DO3A-diph and Gd-AAZTAC17, respectively. Their LDL adducts are taken up by human hepatoblastoma G2 (HepG2 and melanoma B16 tumor cells when added to the incubation medium. T, measurements of the labeled cells indicate that Gd-AAZTAC17 is significantly more efficient than Gd-DO3A-diph. Furthermore, it has been found that HepG2 hepatoma cells can internalize higher amounts of Gd-AAZTAC17 than B16 cells and the involvement of LDL receptors (LDLRs has been demonstrated in competition assays with free LDL. Gd-AAZTAC17/LDL adduct proved to be an efficient probe in the magnetic resonance (MR visualization of subcutaneous tumors in animal models obtained by injecting B16 melanoma cells into the right flank of mice. Finally, confocal microscopy validation of the distribution of LDL-based probes in the tumor has been obtained by doping the Gd-AAZTAC17/LDL adduct with a fluorescent phospholipid moiety.

  19. 3D characterization of EMT cell density in developing cardiac cushions using optical coherence tomography (Conference Presentation)

    Science.gov (United States)

    Yu, Siyao; Gu, Shi; Zhao, Xiaowei; Liu, Yehe; Jenkins, Michael W.; Watanabe, Michiko; Rollins, Andrew M.

    2017-02-01

    Congenital heart defects (CHDs) are the most common birth defect, affecting between 4 and 75 per 1,000 live births depending on the inclusion criteria. Many of these defects can be traced to defects of cardiac cushions, critical structures during development that serve as precursors to many structures in the mature heart, including the atrial and ventricular septa, and all four sets of cardiac valves. Epithelial-mesenchymal transition (EMT) is the process through which cardiac cushions become populated with cells. Altered cushion size or altered cushion cell density has been linked to many forms of CHDs, however, quantitation of cell density in the complex 3D cushion structure poses a significant challenge to conventional histology. Optical coherence tomography (OCT) is a technique capable of 3D imaging of the developing heart, but typically lacks the resolution to differentiate individual cells. Our goal is to develop an algorithm to quantitatively characterize the density of cells in the developing cushion using 3D OCT imaging. First, in a heart volume, the atrioventricular (AV) cushions were manually segmented. Next, all voxel values in the region of interest were pooled together to generate a histogram. Finally, two populations of voxels were classified using either K-means classification, or a Gaussian mixture model (GMM). The voxel population with higher values represents cells in the cushion. To test the algorithm, we imaged and evaluated avian embryonic hearts at looping stages. As expected, our result suggested that the cell density increases with developmental stages. We validated the technique against scoring by expert readers.

  20. Monocyte-Derived Dendritic Cells Upregulate Extracellular Catabolism of Aggregated Low-Density Lipoprotein on Maturation, Leading to Foam Cell Formation.

    Science.gov (United States)

    Haka, Abigail S; Singh, Rajesh K; Grosheva, Inna; Hoffner, Haley; Capetillo-Zarate, Estibaliz; Chin, Harvey F; Anandasabapathy, Niroshana; Maxfield, Frederick R

    2015-10-01

    Although dendritic cells are known to play a role in atherosclerosis, few studies have examined the contribution of the wide variety of dendritic cell subsets. Accordingly, their roles in atherogenesis remain largely unknown. We investigated the ability of different dendritic cell subsets to become foam cells after contact with aggregated low-density lipoprotein (LDL; the predominant form of LDL found in atherosclerotic plaques). We demonstrate that both murine and human monocyte-derived dendritic cells use exophagy to degrade aggregated LDL, leading to foam cell formation, whereas monocyte-independent dendritic cells are unable to clear LDL aggregates by this mechanism. Exophagy is a catabolic process in which objects that cannot be internalized by phagocytosis (because of their size or association with extracellular structures) are initially digested in an extracellular acidic lytic compartment. Surprisingly, we found that monocyte-derived dendritic cells upregulate exophagy on maturation. This contrasts various forms of endocytic internalization in dendritic cells, which decrease on maturation. Finally, we show that our in vitro results are consistent with dendritic cell lipid accumulation in plaques of an ApoE(-/-) mouse model of atherosclerosis. Our results show that monocyte-derived dendritic cells use exophagy to degrade aggregated LDL and become foam cells, whereas monocyte-independent dendritic cells are unable to clear LDL deposits. Furthermore, we find that exophagy is upregulated on dendritic cell maturation. Thus, exophagy-mediated foam cell formation in monocyte-derived dendritic cells could play a significant role in atherogenesis. © 2015 American Heart Association, Inc.

  1. Glass bead cultivation of fungi

    DEFF Research Database (Denmark)

    Droce, Aida; Sørensen, Jens Laurids; Giese, H.

    2013-01-01

    Production of bioactive compounds and enzymes from filamentous fungi is highly dependent on cultivation conditions. Here we present an easy way to cultivate filamentous fungi on glass beads that allow complete control of nutrient supply. Secondary metabolite production in Fusarium graminearum...... and Fusarium solani cultivated on agar plates, in shaking liquid culture or on glass beads was compared. Agar plate culture and glass bead cultivation yielded comparable results while liquid culture had lower production of secondary metabolites. RNA extraction from glass beads and liquid cultures was easier...... to specific nutrient factors. •Fungal growth on glass beads eases and improves fungal RNA extraction....

  2. The Cultivated Mind: From Mental Mediation to Cultivation.

    Science.gov (United States)

    Josephs, Ingrid E.; Fuhrer, Urs

    1998-01-01

    Examines Simmel's principle of cultivation whereby the cultivated mind is constructed through ongoing transactions of people with their cultural environment, cultural forms currently overlooked. Cultural forms result from externalizations of former person-culture transactions. Argues that development is structured through person-culture…

  3. Plant growth and cultivation.

    Science.gov (United States)

    Podar, Dorina

    2013-01-01

    There is a variety of methods used for growing plants indoor for laboratory research. In most cases plant research requires germination and growth of plants. Often, people have adapted plant cultivation protocols to the conditions and materials at hand in their own laboratory and growth facilities. Here I will provide a guide for growing some of the most frequently used plant species for research, i.e., Arabidopsis thaliana, barley (Hordeum vulgare) and rice (Oryza sativa). However, the methods presented can be used for other plant species as well, especially if they are related to the above-mentioned species. The presented methods include growing plants in soil, hydroponics, and in vitro on plates. This guide is intended as a starting point for those who are just beginning to work on any of the above-mentioned plant species. Methods presented are to be taken as suggestive and modification can be made according to the conditions existing in the host laboratory.

  4. NG2 Proteoglycan Ablation Reduces Foam Cell Formation and Atherogenesis via Decreased Low-Density Lipoprotein Retention by Synthetic Smooth Muscle Cells.

    Science.gov (United States)

    She, Zhi-Gang; Chang, Yunchao; Pang, Hong-Bo; Han, Wenlong; Chen, Hou-Zao; Smith, Jeffrey W; Stallcup, William B

    2016-01-01

    Obesity and hyperlipidemia are critical risk factors for atherosclerosis. Because ablation of NG2 proteoglycan in mice leads to hyperlipidemia and obesity, we investigated the impact of NG2 ablation on atherosclerosis in apoE null mice. Immunostaining indicates that NG2 expression in plaque, primarily by synthetic smooth muscle cells, increases during atherogenesis. NG2 ablation unexpectedly results in decreased (30%) plaque development, despite aggravated obesity and hyperlipidemia. Mechanistic studies reveal that NG2-positive plaque synthetic smooth muscle cells in culture can sequester low-density lipoprotein to enhance foam-cell formation, processes in which NG2 itself plays direct roles. In agreement with these observations, low-density lipoprotein retention and lipid accumulation in the NG2/ApoE knockout aorta is 30% less than that seen in the control aorta. These results indicate that synthetic smooth muscle cell-dependent low-density lipoprotein retention and foam cell formation outweigh obesity and hyperlipidemia in promoting mouse atherogenesis. Our study sheds new light on the role of synthetic smooth muscle cells during atherogenesis. Blocking plaque NG2 or altering synthetic smooth muscle cells function may be promising therapeutic strategies for atherosclerosis. © 2015 American Heart Association, Inc.

  5. Starting from grape cultivation.

    Science.gov (United States)

    Yoshida, A

    1992-06-01

    Rapid population growth can only be stopped by lowering the fertility rate. The UNFPA recommends improving the employment opportunities for women as the single best way of achieving this reduction. An example of this phenomenon is the grape cultivation in the Nordeste (Northeastern) region of Brazil. This area is the poorest part of Brazil and has the highest proportion of indigent people. These people have been deforesting the Amazon in search of a better life. What they have done is sterilize the land and turned a tropical rain forest into a desert. In an effort to reverse this trend, grape cultivation has been introduced in an area called Petrolina. The area is very dry with less than 500 mm of precipitation annually. They do have access to a 5000 square kilometer artificial lake (the largest in the world) and the 3rd largest river in Brazil (the Sao Francisco). In an effort to avoid using agricultural medicines, the vines are fertilized with organic matter created on the farm and little or no pesticides are used since pests do not live in such an arid region. It has taken 20 years of trial and error, but the quality of the grapes is now very high and is competitive on the world market. Because of climate and location, harvesting is done year round which increases the productivity of the land. The farm managers have found that married women make the best workers and have the highest level of productivity. Age at 1st marriage averages 24-25, compared with 15-16 for unemployed women in the same area. The fertility rate averages 50% of that for unemployed women in the same area. Agricultural development offers the best opportunity for the women of developing countries. It can pay a high wage, reduce fertility, and replant desert areas.

  6. Three-dimensional printing of stem cell-laden hydrogels submerged in a hydrophobic high-density fluid.

    Science.gov (United States)

    Duarte Campos, Daniela F; Blaeser, Andreas; Weber, Michael; Jäkel, Jörg; Neuss, Sabine; Jahnen-Dechent, Wilhelm; Fischer, Horst

    2013-03-01

    Over the last decade, bioprinting technologies have begun providing important tissue engineering strategies for regenerative medicine and organ transplantation. The major drawback of past approaches has been poor or inadequate material-printing device and substrate combinations, as well as the relatively small size of the printed construct. Here, we hypothesise that cell-laden hydrogels can be printed when submerged in perfluorotributylamine (C(12)F(27)N), a hydrophobic high-density fluid, and that these cells placed within three-dimensional constructs remain viable allowing for cell proliferation and production of extracellular matrix. Human mesenchymal stem cells and MG-63 cells were encapsulated into agarose hydrogels, and subsequently printed in high aspect ratio in three dimensional structures that were supported in high density fluorocarbon. Three-dimensional structures with various shapes and sizes were manufactured and remained stable for more than six months. Live/dead and DAPI stainings showed viable cells 24 h after the printing process, as well as after 21 days in culture. Histological and immunohistochemical analyses after 14 and 21 days revealed viable cells with marked matrix production and signs of proliferation. The compressive strength values of the printed gels consequently increased during the two weeks in culture, revealing encouraging results for future applications in regenerative medicine.

  7. Long-term cultivation of in vitro Apis mellifera cells by gene transfer of human c-myc proto-oncogene.

    Science.gov (United States)

    Kitagishi, Yasuko; Okumura, Naoko; Yoshida, Hitomi; Nishimura, Yuri; Takahashi, Jun-ichi; Matsuda, Satoru

    2011-08-01

    Establishment of cell lines representative of honeybee character would greatly assist in their analysis. Here, we show that immortalized cell line, designated as MYN9, has been generated from honeybee embryo by the gene transfer of human c-myc proto-oncogene. The morphology of the cell is characteristic of embryonic stem cell, although the cell is stable and does not spontaneously differentiate. Polymerase chain reaction analyses show that the cell is originated from authentic honeybee cell. It is proposed that the integration of human c-myc gene into honeybee precursor populations results in the establishment of stable cell line suitable for cellular and molecular studies.

  8. Hybrid origins of cultivated potatoes

    Science.gov (United States)

    Wild and cultivated potatoes, Solanum section Petota, is taxonomically difficult, partly because of interspecific hybridization at both the diploid and polyploid levels. The taxonomy of cultivated potatoes is particularly controversial. With DNA sequence data of the GBSSI (waxy) gene we here infer r...

  9. "Logistic analysis of algae cultivation"

    NARCIS (Netherlands)

    Slegers, P.M.; Leduc, S.; Wijffels, R.H.; Straten, van G.; Boxtel, van A.J.B.

    2015-01-01

    Energy requirements for resource transport of algae cultivation are unknown. This work describes the quantitative analysis of energy requirements for water and CO2 transport. Algae cultivation models were combined with the quantitative logistic decision model ‘BeWhere’ for the regions Benelux

  10. Differences in infectivity between endosymbiotic Chlorella variabilis cultivated outside host Paramecium bursaria for 50 years and those immediately isolated from host cells after one year of reendosymbiosis

    Directory of Open Access Journals (Sweden)

    Y. Kodama

    2016-01-01

    Full Text Available Chlorella variabilis strain NC64A is an intracellular photobiont of the ciliate Paramecium bursaria. NC64A was isolated from P. bursaria nearly 50 years ago and was thereafter cultivated outside the host. This study was undertaken to detect changes in its infectivity to P. bursaria and its auxotrophy for growth outside the host induced during long-term cultivation. NC64A can grow in Modified Bold's Basal Medium but not in C medium, whereas another symbiotic Chlorella variabilis strain, 1N, that was recently isolated from the host grew in C medium but not in Modified Bold's Basal Medium. With regards infectivity, NC64A in the logarithmic phase of growth showed low infectivity to alga-removed P. bursaria cells, whereas those in the early stationary phase showed high infectivity of about 30%. Those in the decay phase of growth showed no infectivity. Results show that NC64A has infectivity, but the infection rate depends on their culture age in the growth curve. Furthermore, NC64A that had been re-infected to P. bursaria for more than one year and isolated from the host showed a nearly 100% infection rate, which indicates that NC64A can recover its infectivity by re-infection to P. bursaria.

  11. Densidade básica da madeira de sete espécies e três clones de eucalipto antes e durante o cultivo de shiitake = Basic density of wood from seven species and three clones of eucalyptus before and during shiitake cultivation

    Directory of Open Access Journals (Sweden)

    Meire Cristina Nogueira de Andrade

    2009-04-01

    Full Text Available Avaliou-se a densidade básica da madeira e casca de sete espécies (E. saligna, E. grandis, E. urophylla, E. camaldulensis, E. citriodora, E. paniculata e E. pellita e três clones de eucalipto (híbridos de E. grandis x E. urophylla antes e durante o cultivo das linhagens LE-95/01 e LE-96/18 de shiitake (Lentinula edodes em toras. Cada linhagem de shiitake foi inoculada em nove toras de cada tipo de eucalipto com 1 m de comprimento e 9 a 14 cm de diâmetro. Assim, o delineamento experimental foi inteiramente casualizado, com 20tratamentos e 9 repetições, sendo cada repetição correspondente a uma tora. As toras foram mantidas em estufa climatizada, com temperatura de 25 ± 5ºC e umidade relativa do ar entre 60-80% durante 12 meses. Para a determinação da densidade básica, analisaram-secunhas de discos e cascas de eucalipto recém-cortadas (sem inoculação das linhagens de L. edodes e em cunhas de discos retirados de toras já inoculadas com as linhagens de L. edodes após 8 e 12 meses de incubação. Verificou-se que a densidade básica da madeira, aolongo do ciclo de cultivo, foi reduzida em todos os tipos de eucalipto.Basic density of the wood and bark of seven eucalyptus species (E. saligna, E. grandis, E. urophylla, E. camaldulensis, E. citriodora, E.paniculata and E. pellita and three eucalyptus clones (E. grandis x E. urophylla hybrids were evaluated before and during the cultivation of shiitake (Lentinula edodes strains LE-95/01and LE-96/18 in logs. Each shiitake strain was inoculated into 9 logs of each type of eucalyptus with length of 1 m and diameter of 9 to 14 cm. Thus, the experimental design was totally randomized, with 20 treatments and 9 repetitions, with each repetition corresponding to one log. The logs were kept in a greenhouse, under the temperature of25 ± 5ºC and relative air humidity between 60-80 %, during 12 months. To determine basic density, newly cut disks and barks wedges of eucalyptus (without the

  12. Densidade de plantas e número de drenos influenciando a produtividade de roseiras cultivadas em vaso Planting density and number of drains influencing the productivity of rose plants cultivated in pots

    Directory of Open Access Journals (Sweden)

    Thales VA Viana

    2008-12-01

    Full Text Available O cultivo de flores no estado do Ceará vem se ampliando nos últimos anos, principalmente nas regiões serranas que proporcionam clima favorável ao desenvolvimento de diversas culturas. Entretanto, poucos são os trabalhos desenvolvidos nessa área, fazendo com que os produtores se utilizem do empirismo no desenvolvimento dos cultivos. Por conseguinte, esse trabalho teve como objetivo avaliar os efeitos do número de plantas (2, 3 e 4 e da quantidade de drenos (1 e 8 por vaso no número de hastes por vaso da roseira, em ambiente protegido. O experimento foi conduzido na Empresa Reijers, no município de São Benedito-CE, sendo o delineamento experimental em blocos casualizados em arranjo fatorial 3 x 2, com quatro repetições. Avaliou-se o número de hastes com 35, 40, 50 e 60 cm e, o número total de hastes por vasos de 12,0 L. As maiores quantidades de plantas por vaso proporcionaram um maior número de hastes por área, mas com predomínio de hastes de menor valor comercial. A utilização de um menor número de plantas por vaso resultou em um menor número de hastes por área, mas em maior número de hastes com maior valor comercial. A utilização de vasos com maior número de drenos reduziu o número total de hastes por vaso.The cultivation of flowers in Ceará State has expanded during the last years, especially in highland areas with a favorable climate for several crops. However, there exist only limited research in this area, so that producers work empirically with those crops. Consequently, this research aimed to evaluate the effects of the number of plants (2, 3 and 4 and the amount of drains (1 and 8 per pot on the number of stems per pot, in a protected environment. The experiment was carried out in São Benedito, Ceará State, Brazil, following a 3 x 2 factorial randomized block design with four repetitions. The number of stems with 35, 40, 50, 60 cm and, the total number of stems per pots of 12.0 L were evaluated. The highest

  13. Density and morphology of corneal endothelial cell after phacoemulsification using Ringer lactate versus balanced salt solution as irrigating solutions

    Directory of Open Access Journals (Sweden)

    Farahdina Rahmawati

    2018-02-01

    Full Text Available AIM: To compare the difference in corneal endothelial cell density and morphology after phacoemulsification using Ringer lactate(RLand balanced salt solution(BSSirrigating solutions.METHODS: The prospective randomized controlled trial study was conducted between February 2017 and April 2017 in Dr. YAP Eye Hospital, Yogyakarta, Indonesia. There were a total of 52 subjects(52 eyeswho were senile cataract patients further grouped into two, 26 patients undergoing the phacoemulsification procedure using RL irrigating solution and the other 26 patients with BSS irrigating solution, both conducted by one operator. On the 1, 7, and 28d post operative, an evaluation was done to measure the density and corneal endothelial cell morphology, as well as the variable of inflammation in the two groups.RESULTS: Fifty-two eyes had undergone phacoemulsification with posterior intraocular lens implantation. Both groups were evaluated for the endothelial cell reduction and corneal endothelial cell morphology change, along with post-operative inflammation. On the 28d post-operative, endothelial cell reduction in the BSS group(173.96 cell/mm2, 8.12%was lower than the RL group(253.20 cell/mm2, 10.25%, percentage of corneal endothelial cell variation coefficient increase in the BSS group(2.92%, 8.36%was lower compared to the RL group(3.42%, 9.96%, decrease of hexagonal cells of corneal endothelium cells presentation percentage in the BSS group(4.30%, 8.17%was lower compared to the RL group(4.84%, 8.97%, and the percentage increase of central corneal thickness in the BSS group(4.69 μm, 0.89%was almost equal to the RL group(4.53 μm, 0.90%. All of the results regarding difference in density and corneal cell endothelium morphology between the two groups did not reveal any statistically significant difference(P>0.05. Inflammatory variable in the two groups were even.CONCLUSION: BSS and RL were equal in their capability of maintaining endothelial cell loss and endothelial

  14. A Breast Cell Atlas: Organelle analysis of the MDA-MB-231 cell line by density-gradient fractionation using isotopic marking and label-free analysis

    Directory of Open Access Journals (Sweden)

    Marianne Sandin

    2015-09-01

    Full Text Available Protein translocation between organelles in the cell is an important process that regulates many cellular functions. However, organelles can rarely be isolated to purity so several methods have been developed to analyse the fractions obtained by density gradient centrifugation. We present an analysis of the distribution of proteins amongst organelles in the human breast cell line, MDA-MB-231 using two approaches: an isotopic labelling and a label-free approach.

  15. The role of cell body density in ruminant retina mechanics assessed by atomic force and Brillouin microscopy.

    Science.gov (United States)

    Weber, Isabell; Yun, Seok-Hyun; Scarcelli, Guillano; Franze, Kristian

    2017-04-13

    Cells in the central nervous system (CNS) respond to the stiffness of their environment. CNS tissue is mechanically highly heterogeneous, thus providing motile cells with region-specific mechanical signals. While CNS mechanics has been measured with a variety of techniques, reported values of tissue stiffness vary greatly, and the morphological structures underlying spatial changes in tissue stiffness remain poorly understood. We here exploited two complementary techniques, contact-based atomic force microscopy and contact-free Brillouin microscopy, to determine the mechanical properties of ruminant retinae, which are built up by different tissue layers. As in all vertebrate retinae, layers of high cell body densities ('nuclear layers') alternate with layers of low cell body densities ('plexiform layers'). Different tissue layers varied significantly in their mechanical properties, with the photoreceptor layer being the stiffest region of the retina, and the inner plexiform layer belonging to the softest regions. As both techniques yielded similar results, our measurements allowed us to calibrate the Brillouin microscopy measurements and convert the Brillouin shift into a quantitative assessment of elastic tissue stiffness with optical resolution. Similar as in the mouse spinal cord and the developing Xenopus brain, we found a strong correlation between nuclear densities and tissue stiffness. Hence, the cellular composition of retinae appears to strongly contribute to local tissue stiffness, and Brillouin microscopy shows a great potential for the application in vivo to measure the mechanical properties of transparent tissues. © 2017 IOP Publishing Ltd.

  16. The role of cell body density in ruminant retina mechanics assessed by atomic force and Brillouin microscopy

    Science.gov (United States)

    Weber, Isabell P.; Yun, Seok Hyun; Scarcelli, Giuliano; Franze, Kristian

    2017-12-01

    Cells in the central nervous system (CNS) respond to the stiffness of their environment. CNS tissue is mechanically highly heterogeneous, thus providing motile cells with region-specific mechanical signals. While CNS mechanics has been measured with a variety of techniques, reported values of tissue stiffness vary greatly, and the morphological structures underlying spatial changes in tissue stiffness remain poorly understood. We here exploited two complementary techniques, contact-based atomic force microscopy and contact-free Brillouin microscopy, to determine the mechanical properties of ruminant retinae, which are built up by different tissue layers. As in all vertebrate retinae, layers of high cell body densities (‘nuclear layers’) alternate with layers of low cell body densities (‘plexiform layers’). Different tissue layers varied significantly in their mechanical properties, with the photoreceptor layer being the stiffest region of the retina, and the inner plexiform layer belonging to the softest regions. As both techniques yielded similar results, our measurements allowed us to calibrate the Brillouin microscopy measurements and convert the Brillouin shift into a quantitative assessment of elastic tissue stiffness with optical resolution. Similar as in the mouse spinal cord and the developing Xenopus brain, we found a strong correlation between nuclear densities and tissue stiffness. Hence, the cellular composition of retinae appears to strongly contribute to local tissue stiffness, and Brillouin microscopy shows a great potential for the application in vivo to measure the mechanical properties of transparent tissues.

  17. Cytotoxic activities of 9,11-dehydroergosterol peroxide and ergosterol peroxide from the fermentation mycelia of ganoderma lucidum cultivated in the medium containing leguminous plants on Hep 3B cells.

    Science.gov (United States)

    Chen, Yu-Kuo; Kuo, Yueh-Hsiung; Chiang, Been-Huang; Lo, Jir-Mehng; Sheen, Lee-Yan

    2009-07-08

    The objective of this study was to investigate the cytotoxicity of the ethanolic extract of mycelia from Ganoderma lucidum (EMG) cultivated in a medium containing leguminous plants Glycine max (L.) Merr. and Astragalus membranaceus on human hepatocellular carcinoma cells (Hep 3B) and to isolate the active components from EMG. The results indicated that EMG induced cytotoxicity in a dose- and time-dependent manner, and the cells treated with EMG for 24, 48, and 72 h had IC(50) values of 156.8, 89.9, and 70.1 microg/mL, respectively. Furthermore, EMG was fractionated into seven fractions (F1-F7). We found that F5 and F6 had higher growth inhibitory effects on Hep 3B cells than the other fractions, and F6 possessed enough amounts (about 2.1 g) to carry out a more detailed study. F6 caused a sub-G1 peak rise and DNA fragmentation in Hep 3B cells and was further separated by high-performance liquid chromatography to obtain two active compounds, 9,11-dehydroergosterol peroxide [9(11)-DHEP] (compound 1) and ergosterol peroxide (EP) (compound 2). The IC(50) values of 9(11)-DHEP and EP based on the cell viability of Hep 3B were 16.7 and 19.4 microg/mL, respectively.

  18. Viabilidade econômica do cultivo de carpa comum (cyprinus carpio linnaeus, 1758 em monocultivo em densidades diferentes Economical viability of the common carp (Cyprinus carpio Linnaeus, 1758 cultivation in monocultivations in different densities

    Directory of Open Access Journals (Sweden)

    Álvaro Graeff

    2004-06-01

    Full Text Available Objetivou-se com esta pesquisa estudar o efeito econômico do povoamento de alevinos I de carpa comum (Cyprinus carpio LINNAEUS, 1758 com densidade diferenciada e com alimentação artificial e adubação química. O delineamento foi inteiramente casualizado com três tratamentos (1; 0,5 e 0,33 peixes/m² avaliados ao longo do tempo e três repetições. Os peixes foram estocados com peso e comprimento médios de 2,40 g e 5,35; 5,40 e 5,39 cm, respectivamente, nos tratamentos, e com 45 dias de idade. As dietas foram formuladas com 27% de PB e 2925 Kcal/Kg de ração, sendo fornecidas diariamente na quantidade de 3% do peso vivo. Os resultados para o comprimento e peso dos peixes foram 25,68; 28,66 e 31,21 cm e 255,00; 424,00 e 519,66 g, respectivamente. A analise econômica foi estudada por meio de planilha de custo. Na produção total, os resultados foram 1882; 1513 e 1367 kg/hectare em um ciclo. Em condições similares, pode-se recomendar zootecnicamente todas as densidades estudadas e, economicamente, a lucratividade da atividade é crescente com o aumento da densidade, havendo diminuição do custo médio ou unitário.This research aimed at studying the economical effect of common carp (Cyprinus carpio LINNAEUS, 1758 settling of fingerlings with differenciated density and with artificial feeding and chemical fertilization. The design was entirely randomized with three treatments (1; 0.5 and 0.33 fishes/m² evaluated along the time and 3 repetitions. The fish were stocked with medium weight and mediun length of 2.40 g and 5.35; 5.40 and 5.39 cm, respectively, in the treatments, and 45 days old. The diets were formulated with 27% PB and 2.925 kcal/kg of ration, being given daily in the quantities of 3% of living weight. The results for the length and weight of the fishes were 25.68; 28.66 and 31.21 cm and 255.00; 424.00 and 519.66 g, respectively. The economical analisys was studied through cost planning. In the total production, the results

  19. Dissecting the actin cortex density and membrane-cortex distance in living cells by super-resolution microscopy

    DEFF Research Database (Denmark)

    Clausen, M. P.; Colin-York, H.; Schneider, Falk

    2017-01-01

    and accurately measure the density distribution of the cortical actin cytoskeleton and the distance between the actin cortex and the membrane in live Jurkat T-cells. We found an asymmetric cortical actin density distribution with a mean width of 230 (+105/-125) nm. The spatial distances measured between......Nanoscale spacing between the plasma membrane and the underlying cortical actin cytoskeleton profoundly modulates cellular morphology, mechanics, and function. Measuring this distance has been a key challenge in cell biology. Current methods for dissecting the nanoscale spacing either limit...... themselves to complex survey design using fixed samples or rely on diffraction-limited fluorescence imaging whose spatial resolution is insufficient to quantify distances on the nanoscale. Using dual-color super-resolution STED (stimulated-emission-depletion) microscopy, we here overcome this challenge...

  20. The role of surface charge density in cationic liposome-promoted dendritic cell maturation and vaccine-induced immune responses

    Science.gov (United States)

    Ma, Yifan; Zhuang, Yan; Xie, Xiaofang; Wang, Ce; Wang, Fei; Zhou, Dongmei; Zeng, Jianqiang; Cai, Lintao

    2011-05-01

    Cationic liposomes have emerged as a novel adjuvant and antigen delivery system to enhance vaccine efficacy. However, the role of surface charge density in cationic liposome-regulated immune responses has not yet been elucidated. In the present study, we prepared a series of DOTAP/DOPC cationic liposomes with different surface densities by incorporating varying amounts of DOPC (a neutral lipid) into DOTAP (a cationic lipid). The results showed that DOTAP/DOPC cationic liposome-regulated immune responses relied on the surface charge density, and might occur through ROS signaling. The liposomes with a relatively high charge density, such as DOTAP/DOPC 5 : 0 and 4 : 1 liposomes, potently enhanced dendritic cell maturation, ROS generaion, antigen uptake, as well as the production of OVA-specific IgG2a and IFN-γ. In contrast, low-charge liposomes, such as DOTAP/DOPC 1 : 4 liposome, failed to promote immune responses even at high concentrations, confirming that the immunoregulatory effect of cationic liposomes is mostly attributable to their surface charge density. Moreover, the DOTAP/DOPC 1 : 4 liposome suppressed anti-OVA antibody responses in vivo. Overall, maintaining an appropriate surface charge is crucial for optimizing the adjuvant effect of cationic liposomes and enhancing the efficacy of liposome-based vaccines.

  1. Morphology and rheology in filamentous cultivations.

    Science.gov (United States)

    Wucherpfennig, T; Kiep, K A; Driouch, H; Wittmann, C; Krull, R

    2010-01-01

    Because of their metabolic diversity, high production capacity, secretion efficiency, and capability of carrying out posttranslational modifications, filamentous fungi are widely exploited as efficient cell factories in the production of metabolites, bioactive substances, and native or heterologous proteins, respectively. There is, however, a complex relationship between the morphology of these microorganisms, transport phenomena, the viscosity of the cultivation broth, and related productivity. The morphological characteristics vary between freely dispersed mycelia and distinct pellets of aggregated biomass, every growth form having a distinct influence on broth rheology. Hence, the advantages and disadvantages for mycelial or pellet cultivation have to be balanced out carefully. Because of the still inadequate understanding of the morphogenesis of filamentous microorganisms, fungal morphology is often a bottleneck of productivity in industrial production. To obtain an optimized production process, it is of great importance to gain a better understanding of the molecular and cell biology of these microorganisms as well as the relevant approaches in biochemical engineering. In this chapter, morphology and growth of filamentous fungi are described, with special attention given to specific problems as they arise from fungal growth forms; growth and mass transfer in fungal biopellets are discussed as an example. To emphasize the importance of the flow behavior of filamentous cultivation broths, an introduction to rheology is also given, reviewing important rheological models and recent studies concerning rheological parameters. Furthermore, current knowledge on morphology and productivity in relation to the environom is outlined in the last section of this review. Copyright 2010 Elsevier Inc. All rights reserved.

  2. Processing of acetylated human low-density lipoprotein by parenchymal and non-parenchymal liver cells. Involvement of calmodulin?

    OpenAIRE

    Van Berkel, Theo J.C.; Nagelkerke, Jan F.; Harkes, Leen; Kruijt, Johan K.

    1982-01-01

    1. Modified lipoproteins have been implicated to play a significant role in the pathogenesis of atherosclerosis. In view of this we studied the fate and mechanism of uptake in vivo of acetylated human low-density lipoprotein (acetyl-LDL). Injected intravenously into rats, acetyl-LDL is rapidly cleared from the blood. At 10min after intravenous injection, 83% of the injected dose is recovered in liver. Separation of the liver into a parenchymal and non-parenchymal cell fraction indicates that ...

  3. Distribution and density of CD1a+ and CD83+ dendritic cells in HPV-associated laryngeal papillomas.

    Science.gov (United States)

    Kovalenko, Sergey; Lukashenko, Polina; Romanovskaya, Alesia; Soldatski, Iouri L; Bakanov, Sergey I; Pfister, Herbert; Gerein, Valentin

    2009-02-01

    Respiratory papillomatosis associated with human papilloma virus (HPV) infection is the most common benign laryngeal neoplasm. The age of patients at disease onset, HPV type, number of surgeries are well known prognostic factors of the disease course. The correlation between dendritic cell (DC) density in tumor tissue and clinical prognosis was established. The aim of our study was to estimate the density of DC in laryngeal papillomas associated with HPV types 6/11 infection and to evaluate the relationship between the number of DC and the disease severity. Our study included 40 randomly selected biopsy specimens from patients with HPV-positive laryngeal papillomatosis aged from 1.7 to 20 year. DC were immunohistochemically labelled with anti-CD1a antibodies and anti-CD83 antibodies. The density of DC was analysed in epithelial layer and lamina propria. In the epithelial layer of papillomas the number of CD1a+ and CD83+ DC was 86.2 (47.5-119.9) cells/mm(2) and 2.6 (0.6-7.9) cells/mm(2), respectively. In lamina propria - 15.3 (5.1-27.9) and 16.0 (6.7-33.2) cells/mm(2). For subgroups of patients with high number of operations (more than 3), early disease onset (children under 3 years of age) and lingering duration of disease (more than 1 year) we detected an increase of CD83+ DC in the epithelial layer. However, our data did not demonstrate a statistically significant difference in CD1a+ DC count neither in the epithelium nor in the lamina propria. Probably, the increase of CD83+ DC density in epithelial layer of patients with severe course of disease can be an evidence of impaired migration of matured DC.

  4. In vitro cultivation of Maritrema novaezealandensis (Microphallidae)

    DEFF Research Database (Denmark)

    Fredensborg, Brian Lund; Poulin, R

    2005-01-01

    In this study, the effect of the composition of culture medium on excystation, growth, survival and egg production was investigated for the recently discovered microphallid trematode Maritrema novaezealandensis. Metacercariae from the two second intermediate crab hosts, Macrophthalmus hirtipes...... and Halicarcinus whitei, were incubated in either: (1) 0.85% saline solution, (2) the commercial cell culture medium, NCTC-109, (3) NCTC-109 supplemented with 20%, or (4) NCTC-109 supplemented with 40% chicken serum. Furthermore, excysted metacercariae were cultured for 5 days in each of the three media: NCTC-109...... cultured in media with a supplement of chicken serum and reached a maximum after 2 days of cultivation. Growth, however, did not occur after the first day of cultivation in any of the three media....

  5. High energy efficiency and high power density proton exchange membrane fuel cells: Electrode kinetics and mass transport

    Science.gov (United States)

    Srinivasan, Supramaniam; Velev, Omourtag A.; Parthasathy, Arvind; Manko, David J.; Appleby, A. John

    1991-01-01

    The development of proton exchange membrane (PEM) fuel cell power plants with high energy efficiencies and high power densities is gaining momentum because of the vital need of such high levels of performance for extraterrestrial (space, underwater) and terrestrial (power source for electric vehicles) applications. Since 1987, considerable progress has been made in achieving energy efficiencies of about 60 percent at a current density of 200 mA/sq cm and high power densities (greater than 1 W/sq cm) in PEM fuel cells with high (4 mg/sq cm) or low (0.4 mg/sq cm) platinum loadings in electrodes. The following areas are discussed: (1) methods to obtain these high levels of performance with low Pt loading electrodes - by proton conductor impregnation into electrodes, localization of Pt near front surface; (2) a novel microelectrode technique which yields electrode kinetic parameters for oxygen reduction and mass transport parameters; (3) demonstration of lack of water transport from anode to cathode; (4) modeling analysis of PEM fuel cell for comparison with experimental results and predicting further improvements in performance; and (5) recommendations of needed research and development for achieving the above goals.

  6. [Effect of drugs with acid-neutralizing and acid-suppressive effect on parietal cell function and on gastrin and somatostatin cell density of the stomach].

    Science.gov (United States)

    Arnold, R; Koop, H; Nesslinger, A; Schwarting, H

    1983-03-01

    4 weeks treatment of healthy volunteers with Magaldrate (Al-Mg-containing antacid) is followed by an increase of basal serum gastrin, and a decrease of the antral G-cell density. Gastric acid secretion remained unchanged. In contrast, during maintenance therapy of duodenal ulcer patients with 400 mg cimetidine an increase of the postprandial gastrin output can be observed after 12 months treatment, whereas basal serum gastrin, antral G-cell density and gastric acid secretion remained unchanged. These results suggest that the effects observed during antacid treatment can not be related to an increase in gastric pH and may rather reflect a pH independent direct action of Magaldrate on the antral G-cell.

  7. The effect of cultivation media and washing whole-cell biocatalysts on monoamine oxidase catalyzed oxidative desymmetrization of 3-azabicyclo[3,3,0]octane

    DEFF Research Database (Denmark)

    Ramesh, Hemalata; Zajkoska, Petra; Rebros, Martin

    2016-01-01

    It is well known that washing whole-cells containing enzyme activities after fermentation, but prior to biocatalysis can improve their activity in the subsequent reaction. In this paper, we quantify the impact of both the fermentation media and cell washing on the performance of whole-cell biocat...

  8. Low Temperature, High Energy Density Micro Thin Film Solid Oxide Fuel Cell Project

    Data.gov (United States)

    National Aeronautics and Space Administration — A new type of solid oxide fuel cell based on thin film technology and ultra-thin electrolyte is being proposed to develop to realize major reductions in fuel cell...

  9. Progresso da ferrugem do cafeeiro irrigado em diferentes densidades de plantio pós-poda Progress of rust in coffee plants in various densities of cultivation in irrigated planting after pruning

    Directory of Open Access Journals (Sweden)

    Bernardo Reis Teixeira Lacerda Paiva

    2011-02-01

    Full Text Available Objetivou-se, no presente trabalho, avaliar o efeito de diferentes critérios para manejo da irrigação em quatro densidades de plantio, sob sistema de gotejamento na incidência e severidade da ferrugem do cafeeiro e avaliar a influência do enfolhamento na curva de progresso dessa doença. Conduziu-se, o experimento, em área experimental da Universidade Federal de Lavras MG, utilizando a cultivar Rubi MG-1192 com seis anos. O delineamento experimental foi em blocos ao acaso com quatro repetições. Os tratamentos foram constituídos por quatro parcelas representadas pelas densidades de plantio (convencionais e adensados: 2500 (4,0x1,0 m, 3333 (3,0x1,0 m, 5000 (2,0x1,0 m, 10000 (2,0x0,5 m plantas ha-1, quatro subparcelas sendo: irrigações quando a tensão da água no solo atingiu valores de 20 e 60kPa; irrigações utilizando o manejo do balanço hídrico (calculado através do software IRRIPLUS, com turnos de irrigação fixos de três dias por semana e uma testemunha sem irrigação, perfazendo um total de 16 tratamentos. Cada subparcela foi constituída por 10 plantas, sendo consideradas como plantas úteis as seis centrais. Foram avaliadas a incidência e severidade da ferrugem e a porcentagem de enfolhamento das plantas de cafeeiros. Após análise estatística, os dados foram convertidos em área abaixo da curva de progresso da doença e do crescimento. Verificou-se que os critérios para manejo da irrigação influenciaram a curva de progresso do crescimento, porém, não interferiu na curva de progresso da incidência e da severidade da ferrugem. Os sistemas de plantios adensados favoreceram a incidência da ferrugem. Mas as densidades de plantio não interferiram no enfolhamento.The objective of this study was to evaluate the effect of different irrigation controls implemented in four planting densities on a system of drip on the incidence and severity of rust and to assess the influence of leaf growth on the progress curve of this

  10. Mast Cells Density in Fibrotic Capsule of Enchondroma and Well-Differentiated Chondrosarcoma: A Method for Histopathologic Differentiation

    Directory of Open Access Journals (Sweden)

    Mohammad Javad Kharazi Fard

    2012-02-01

    Full Text Available Background: An enchondroma is a benign and a well-differentiated chondrosarcoma is an invasive chondroid tumor with high recurrence potential. In spite of biologic differences, these two tumors have very similar histopathologic appearance. It has been shown that the biologic nature of the connective tissue around benign and malignant tumors varies in the number of mast cells. The aim of this study was to study the histopathologic distinction of enchondroma and well-differentiated chondrosarcoma using the density of the mast cells in fibrotic capsule. Methods: Twelve enchondroma and 15 well-differentiated chondrosarcoma were collected from Pathology department of Cancer Institute and Central Pathology department of Imam Khomeini Hospital in Tehran. 3 micron paraffin embedded tissue sections were stained by toluidine blue for mast cells counting. Mast cells were counted in fibrous capsule of all cases. Mast cells counts