Human erythrocyte flickering: temperature,ATP concentration,water transport,and cell aging,plus a computer simulation |
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Authors: | David Szekely Tsz Wai Yau Philip W Kuchel |
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Institution: | (1) School of Molecular and Microbial Biosciences, University of Sydney, Building G08, Sydney, NSW, 2006, Australia;(2) Centre for Mathematical Biology, University of Sydney, Sydney, NSW, 2006, Australia |
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Abstract: | Images of human erythrocytes from a healthy donor were recorded under differential interference contrast (DIC) microscopy;
they were acquired rapidly (~336 Hz) and the intensity of the centermost pixel of each cell was recorded for ~60 s (20,000
values). Various techniques were used to analyze the data, including detrended fluctuation analysis (DFA) and multiscale entropy
(MSE); however, power spectrum analysis was deemed the most appropriate for metrifying and comparing results. This analysis
was used to compare cells from young and old populations, and after perturbing normal conditions, with changes in temperature,
adenosine triphosphate (ATP) concentration (using NaF, an inhibitor of glycolysis, and α-toxin, a pore-forming molecule used
to permeabilize red cells to ATP), and water transport rates using glycerol, and p-chloromercuriphenylsulfonic acid (pCMBS) to inhibit aquaporins, AQPs]. There were measurable differences in the membrane
fluctuation characteristics in populations of young and old cells, but there was no significant change in the flickering time
series on changing the temperature of an individual cell, by depleting it of ATP, or by competing with the minor water exchange
pathway via AQP3 using glycerol. However, pCMBS, which inhibits AQP1, the major water exchange pathway, inhibited flickering
in all cells, and yet it was restored by the membrane intercalating species dibutyl phthalate (DBP). We developed a computer
model to simulate acquired displacement spectral time courses and to evaluate various methods of data analysis, and showed
how the flexibility of the membrane, as defined in the model, affects the flickering time course. |
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Keywords: | ATP concentration Flickering DIC microscopy High-speed imaging Human red blood cells In vivo aging Membrane undulations Numerical simulation Spectrin cytoskeleton Water transport |
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