15 research outputs found

    Time-averaged Kuramoto order parameter

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    <p><b> for experiments at different cell densities </b><b>.</b> At cell densities above 0.3%, is high indicating coherent oscillatory dynamics of the cells in the population. At cell densities below 0.3% the low value of shows that the cells oscillate in a decoherent fashion. The critical cell density lies at 0.3%. The error bars represent the standard deviation and provide a measure for the fluctuations of the phase synchronisation in each individual experiment.</p

    Collective and individual dynamics of <i>S. carlsbergensis</i> at a cell density of 0.01%.

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    <p>At  = 0 an aliquot of 52 mM glucose was added to the starved and immobilised cells. <i>(a)</i> The time-series of the macroscopic, collective fluorescence signal is noisy but quiescent. Transient reminicences of oscillations can be spotted at 200 s400 s. <i>(b)</i> The individual cells remain oscillatory, even at this low cell density. The normalised oscillation amplitudes of the individual cells are decoherent. The amplitude of the oscillations of the cells corresponds to 120–200 photons/s.) Note, that the time spans where individual cells show pronounced oscillatory amplitudes vary considerably from cell to cell. The cells are numbered in a random order. <i>(c)</i> The plot of the phases of the oscillations shows that each cell oscillates, however, with its own phase and oscillation period. A slight and transient entrainment in the oscillation phases may be observed for some cells, giving rise to the reminicences of oscillations seen at collective signal at 200 s400 s. <i>(d)</i> The time-dependent Kuramoto order parameter remains low at all times, indicating a complete desynchronisation among the oscillations of the individual cells.</p

    Dynamics of a <i>S. carlsbergensis</i> population of very low density (

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    <p><b> = 0.001%).</b> At  = 0 an aliquot of 52 mM glucose was added to the starved and immobilised cells. While the collective signal (black line) is quiescent and noisy, long-lasting oscillations are observed in the fluorescence signal emitted by an individual cell (gray line). The original time series were smoothed by adjacent averaging over three consecutive data points.</p

    Desynchronisation of Glycolytic Oscillations in Yeast Cell Populations

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    <div><p>Glycolytic oscillations of intact yeast cells of the strain <em>Saccharomyces carlsbergensis</em> were investigated at both the levels of cell populations and of individual cells. Individual cells showed glycolytic oscillations even at very low cell densities (e.g. 1.010<sup>5</sup> cells/ml). By contrast, the collective behaviour on the population level was cell density-dependent: at high cell densities it is oscillatory, but below the threshold density of 1.010<sup>6</sup> cells/ml the collective dynamics becomes quiescent. We demonstrate that the transition in the collective dynamics is caused by the desynchronisation of the oscillations of individual cells. This is characteristic for a Kuramoto transition. Spatially resolved measurements at low cell densities revealed that even cells that adhere to their neighbours oscillated with their own, independent frequencies and phases.</p> </div

    Collective and individual dynamics of <i>S. carlsbergensis</i> at a cell density of 0.1%.

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    <p>At  = 0 an aliquot of 52 mM glucose was added to the starved and immobilised cells. <i>(a)</i> The time-series of the macroscopic, collective fluorescence signal is quiescent. At  = 850 s a transient episode of synchronised, collective oscillations sets in. After 1350 s, the collective signal becomes quiescent again. <i>(b)</i> The individual cells remain oscillatory during the entire duration of the experiment. Remarkably, the synchronisation episode sets in as the normalised amplitudes of the glycolytic oscillations already diminishes. (The amplitude of the oscillations of the cells corresponds to 45–95 photons/s.) The cells are numbered in an random order. <i>(c)</i> At the begin and end of the experiment, the individual cells oscillate at their own periods and phases. The synchronisation episode at 850 s1350 s is caused by a temporary entrainment of the oscillations of the individual cells. <i>(d)</i> The time-dependent Kuramoto order parameter reflects the dynamics in the cell population. It remains below 0.3 as the cells oscillate in a incoherent manner, and increases to levels of 0.7 during the synchronisation episode.</p

    Collective and individual dynamics of a population of cells from <i>S. carlsbergensis</i> at a cell density of 0.7%.

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    <p>At  = 0 an aliquot of 52 mM glucose was added to the starved and immobilised cells. During the first 350 s of the time-series, the cells start to oscillate and to synchronise to each other. <i>(a)</i> The time-series of the macroscopic, collective fluorescence signal shows well-developed glycolytic oscillations. This coherent signal is generated by all cells of the population. <i>(b)</i> The oscillations of the individual cells show that their normalised amplitudes are highly coherent and synchronised. (The amplitude of oscillations of the cells corresponds to 60–140 photons/s.) Note that the dynamics of every cell is plotted as a thin line and its corresponding normalised amplitude is colour-coded. Cells are numbered randomly, i.e., the numbering does not reflect any spatial arrangement of the cells. At 1800 s the oscillation amplitudes decay and the cells lose synchrony due to exhaustion of glucose. <i>(c)</i> The phases of the oscillations of the individual cells are highly synchronised to each other. Again, for 1800 s the cells desynchronise. <i>(d)</i> The time-dependent Kuramoto order parameter indicates the degree of synchronisation among the cells. For 350 s1800 s the order parameter is almost 1, indicating a very high coherence in the oscillations of the individual cells.</p

    Width of the distribution of oscillation periods in individual experiments, as measured by the standard deviation

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    <p><b> of the Gaussian fit to the histogram of oscillation periods.</b> For cell densities below  = 0.3%, 2 s pointing at decoherence in oscillation periods. The line at indicates the sampling rate (2 s). All points below  = 2 s originate from coherent oscillations of the individual cells.</p

    Anchorage independent growth.

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    <p>Both cell lines were seeded in “Soft agar” (S, 0.33%) on “Feeder agar” (F, 0.5%) with (w) or without (wo) the application of EGF and ITS. Caco-2 cells were used as positive control. No anchorage independent growth was detected in IPEC-1 and IPEC-J2. On the other hand, Caco-2 showed an anchorage independent growth which did not depend on the additives EGF and ITS (bar = 20 μm). The results represent at least three independent experiments (n = 3).</p

    Analysis of the protein expression of tight junctions and cytoskeleton—IPEC-1.

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    <p>Different proteins of the tight junctions and cytoskeleton were analysed by Western blot and immunofluorescence: CK18, β-actin, ZO-1, occludin, claudin-3 and claudin-4. At day 2 of culture IPEC-1 showed in the Western blots a weak protein expression of all proteins studied. A strong ZO-1 and occludin immunoreactivity was found at the border of the cells. In IPEC-1 ZO-1 showed a low expression, whereas a large amount of occludin was present. The cytoskeleton proteins CK18 and actin were strongly expressed at the border of the IPEC-1 cells. Furthermore, no stress fibres were detected in the cells. Claudin-3 and claudin-4 were observed in the cells. A spot like character of claudin-3 distribution in the area where at least three cells were closely located. Claudin-4 showed an expression at the cell border but also within the cytoplasm. (blue = DAPI; bar = 20 μm)</p

    Analyses of important genes of the metabolism and oxygen-consumption.

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    <p>Important genes of the metabolism were analysed in both cell lines cultured on membranes for 10 days using microarray analyses and qPCR (A). PDHB (pyruvate dehydrognase subunit B) and CYC1 (cyctochrome C) are significantly down-regulated in the microarray and qPCR. SDH (succinate dehydrogenase subunit B) and HIF1a (hypoxia inducible factor 1a) are both significantly up-regulated in the microarray but not in qPCR. Furthermore, oxygen-consumption of both cell lines cultured on dishes or membranes for 10 days was examined. Both cell lines showed a significant higher O<sub>2</sub>-consumption on membranes (IPEC-1: 20.07 nmol/100 000 cells; IPEC-J2: 75.27 nmol/100 000 cells) in comparison to dishes (IPEC-1: 3.18 nmol/100 000 cells; IPEC-J2: 8.18 nmol/100 000 cells). At the same time, a significant higher oxygen-consumption was found in IPEC-J2 in comparison to IPEC1, which was independent of the culture support.</p
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