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.2020 Aug 20;11(16):6914-6920.
doi: 10.1021/acs.jpclett.0c01748. Epub 2020 Aug 11.

Nanoscale Viscosity of Cytoplasm Is Conserved in Human Cell Lines

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Nanoscale Viscosity of Cytoplasm Is Conserved in Human Cell Lines

Karina Kwapiszewska et al. J Phys Chem Lett..

Abstract

Metabolic reactions in living cells are limited by diffusion of reagents in the cytoplasm. Any attempt to quantify the kinetics of biochemical reactions in the cytosol should be preceded by careful measurements of the physical properties of the cellular interior. The cytoplasm is a complex, crowded fluid characterized by effective viscosity dependent on its structure at a nanoscopic length scale. In this work, we present and validate the model describing the cytoplasmic nanoviscosity, based on measurements in seven human cell lines, for nanoprobes ranging in diameters from 1 to 150 nm. Irrespective of cell line origin (epithelial-mesenchymal, cancerous-noncancerous, male-female, young-adult), we obtained a similar dependence of the viscosity on the size of the nanoprobes, with characteristic length-scales of 20 ± 11 nm (hydrodynamic radii of major crowders in the cytoplasm) and 4.6 ± 0.7 nm (radii of intercrowder gaps). Moreover, we revealed that the cytoplasm behaves as a liquid for length scales smaller than 100 nm and as a physical gel for larger length scales.

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Conflict of interest statement

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
Principle of the researchon cytoplasmic nanoviscosity. (I) Assumptionsof the length-scale dependent viscosity (LSDV) model: (Ia) cytoplasmis a complex liquid containing components of various sizes. Thus,diffusion of the probes of different hydrodynamic radii (rp) is hindered by different cytoplasmic obstacles. Inthe result (Ib), effective viscosity (ηeff) probedby tracers of different sizes increase with the size of the tracer.(II) To examine ηeff, fluorescently labeled tracersare introduced to the cytoplasm—the mode of introduction isoptimized for a given probe. (III) Next, FCS measurements are performed:(IIIa) Confocal spot is positioned in the cytoplasmic area of thecell, and fluorescence fluctuations are registered, (IIIb) autocorrelationcurve (ACC) is calculated for the acquired data, and (IIIc) ACC isfitted with a proper diffusion model, and diffusion coefficient ofthe tracer is derived. (IV) Data collected for a set of tracers ina given cell line is used for quantitative description of the LSDVmodel: (IVa) ηeff experienced by the given probeis calculated from the diffusion coefficient, andrp (IVb) results are plotted and fitted with eq 1; (IVc) LSDV profiles are comparedbetween different cell lines.
Figure 2
Figure 2
Nanoviscosity measured in six differentcell lines. Experimentalresults are presented as scatter. Each point represents the averagevalue obtained from at least 10 cells from two independent inoculations.Error bars correspond to standard deviations. Dashed line representsLSDV model (eq 1) fittedto experimental data with the following parameters:A = 1.3 (fixed),RH = 20 ± 11 nm,ξ = 4.6 ± 0.7 nm,a = 0.57 ± 0.14.Shading represents the error of the model calculated using the totaldifferential method.
Figure 3
Figure 3
Comparisonof nanoviscosity in different cell types. Graphs representaverage relative nanoviscosity measured in the cytoplasm of differentcells and plotted against hydrodynamic radii of the tracers probingthe viscosity (data consistent with Figure 2) (a–d) Cell lines used in the studywere divided into groups (seeSI 5), accordingto (a) tissue origin, (b) disease, (c) gender of donor, or (d) ageof donor. No deviations of the viscosity could have been observedbetween these groups. (e) Fibroblasts were the only cell line in whichnanoviscosity was found to differ from the major trend for small probes(rp < 10 nm).
Figure 4
Figure 4
Confocal images of subcellular structures of four celllines: A549,HeLa, U2-Os, and Fibroblasts. (a) Staining of cytoskeletal proteins(actin and tubulin) showed no particular differences between celltypes. (b) Immunostaining of endoplasmic reticulum (ER) revealed ahigh abundance of ER in fibroblasts comparing to three other celllines. Scale bars correspond to 10 μm.
Figure 5
Figure 5
Quantificationof ER abundance in different cell types. (a) Exampleconfocal images of ER in different cells. (b) Average abundance ofER (white pixels) and cytosol (black pixels) in cells of various types.
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