Abstract
Mechanical vibrations affect multiple cell properties, including its diffusivity, entropy, internal content organization, and thus—function. Here, we used Differential Interference Contrast (DIC), confocal, and Total Internal Reflection Fluorescence (TIRF) microscopies to study mechanical vibrations in live (Jurkat) T cells. Vibrations were measured via the motion of intracellular particles and plasma membrane. These vibrations depend on adenosine triphosphate (ATP) consumption and on Myosin II activity. We then used spectral analysis of these vibrations to distinguish the effects of thermal agitation, ATP-dependent mechanical work and cytoskeletal visco-elasticity. Parameters of spectral analyses could be related to mean square displacement (MSD) analyses with specific advantages in characterizing intracellular mechanical work. We identified two spectral ranges where mechanical work dominated vibrations of intracellular components: 0–3 Hz for intracellular particles and the plasma-membrane, and 100–150 Hz for the plasma-membrane. The 0–3 Hz vibrations of the cell membrane that we measured in an experimental model of immune synapse (IS) are expected to affect the IS formation and function in effector cells. It may also facilitate immunological escape of extensively vibrating malignant cells.
Highlights
Mechanical work inside living cells plays a significant role in cell physiology (Huang and Ingber, 2005; Tee et al, 2010)
The cytoskeleton is an elastic mesh (Mizuno et al, 2007), and it transfers those forces to intracellular constituents, e.g., vesicles and organelles that are embedded within or Abbreviations: Differential Interference Contrast (DIC), Differential interference contrast; Total Internal Reflection Fluorescence (TIRF), Total internal reflection fluorescence; adenosine triphosphate (ATP), Adenosine triphosphate; mean square displacement (MSD), Mean square displacement; immune synapse (IS), Immune synapse; plasma membrane (PM), Plasma membrane; T-cell antigen receptor (TCR), T cell receptor; MHC, Major histocompatibility complex; APC, Antigen presenting cell; FBM, Fractional Brownian motion; Random walk on a fractal (RWF), Random walk on fractal; Continues Time Random Walk (CTRW), Continues time random walk; power spectral density (PSD), Power spectral density; Brownian Diffusion (BD), Brownian diffusion; Discrete Fourier Transform (DFT), Discrete Fourier transform; sum of squared errors (SSE), Sum of squared errors; probability density function (PDF), Probability density function; SD, standard deviation; region of interest (ROI), Region of interest; amplitude spectral density (ASD), Amplitude spectral density; PFA, Paraformaldehyde; ANOVA, Analysis of Variance
B and SSE capture better the conditions of low intracellular mechanical work in those ATP depleted cells. These results suggest that ATP-dependent cytoskeletal motion significantly contribute to membrane fluctuations at low (
Summary
Mechanical work inside living cells plays a significant role in cell physiology (Huang and Ingber, 2005; Tee et al, 2010). The cytoskeleton is an elastic mesh (Mizuno et al, 2007), and it transfers those forces to intracellular constituents, e.g., vesicles and organelles that are embedded within or Abbreviations: DIC, Differential interference contrast; TIRF, Total internal reflection fluorescence; ATP, Adenosine triphosphate; MSD, Mean square displacement; IS, Immune synapse; PM, Plasma membrane; TCR, T cell receptor; MHC, Major histocompatibility complex; APC, Antigen presenting cell; FBM, Fractional Brownian motion; RWF, Random walk on fractal; CTRW, Continues time random walk; PSD, Power spectral density; BD, Brownian diffusion; DFT, Discrete Fourier transform; SSE, Sum of squared errors; PDF, Probability density function; SD, standard deviation; ROI, Region of interest; ASD, Amplitude spectral density; PFA, Paraformaldehyde; ANOVA, Analysis of Variance
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