22 research outputs found
Actin and microtubule networks contribute differently to cell response for small and large strains
Cytoskeletal filaments provide cells with mechanical stability and organization. The main key players
are actin filaments and microtubules governing a cell’s response to mechanical stimuli. We
investigated the specific influences of these crucial components by deforming MCF-7 epithelial cells at
small(\u845% deformation) and large strains(>5% deformation). To understand specific contributions
of actin filaments and microtubules, we systematically studied cellular responses after treatment with
cytoskeleton influencing drugs. Quantification with the microfluidic optical stretcher allowed
capturing the relative deformation and relaxation of cells under different conditions. We separated
distinctive deformational and relaxational contributions to cell mechanics for actin and microtubule
networks for two orders of magnitude of drug dosages. Disrupting actin filaments via latrunculin A,
for instance, revealed a strain-independent softening. Stabilizing these filaments by treatment with
jasplakinolide yielded cell softening for small strains but showed no significant change at large strains.
In contrast, cells treated with nocodazole to disrupt microtubules displayed a softening at large strains
but remained unchanged at small strains. Stabilizing microtubules within the cells via paclitaxel
revealed no significant changes for deformations at small strains, but concentration-dependent
impact at large strains. This suggests that for suspended cells, the actin cortex is probed at small strains,
while at larger strains; the whole cell is probed with a significant contribution from the microtubule
Complex thermorheology of living cells
Temperature has a reliable and nearly instantaneous influence onmechanical responses of cells.As recently
published, MCF-10Anormal epithelial breast cells follow the time–temperature superposition (TTS)
principle. Here,wemeasured thermorheological behaviour of eightcommoncell types within
physiologically relevant temperatures and appliedTTS to creep compliance curves.Our results showed that
superposition is not universal and was seen in four of the eight investigated cell types. For the other cell
types, transitions of thermorheological responses were observed at 36 °C.Activation energies (EA)were
calculated for all cell types and ranged between 50 and 150 kJmol−1.The scaling factors of the superposition
of creep curves were used to group the cell lines into three categories. They were dependent on relaxation
processes aswell as structural composition of the cells in response tomechanical load and temperature
increase.This study supports the view that temperature is a vital parameter for comparing cell rheological
data and should be precisely controlledwhen designing experiments
Thermal instability of cell nuclei
DNA is known to be a mechanically and thermally stable structure. In its double
stranded form it is densely packed within the cell nucleus and is thermo-resistant
up to 70 °C. In contrast, we found a sudden loss of cell nuclei integrity at
relatively moderate temperatures ranging from 45 to 55 °C. In our study, suspended
cells held in an optical double beam trap were heated under controlled
conditions while monitoring the nuclear shape. At specific critical temperatures,
an irreversible sudden shape transition of the nuclei was observed. These temperature
induced transitions differ in abundance and intensity for various normal
and cancerous epithelial breast cells, which clearly characterizes different cell
types. Our results show that temperatures slightly higher than physiological
conditions are able to induce instabilities of nuclear structures, eventually
leading to cell death. This is a surprising finding since recent thermorheological
cell studies have shown that cells have a lower viscosity and are thus more
deformable upon temperature increase. Since the nucleus is tightly coupled to
the outer cell shape via the cytoskeleton, the force propagation of nuclear
reshaping to the cell membrane was investigated in combination with the
application of cytoskeletal drugs
Normal epithelial and triple-negative breast cancer cells show the same invasion potential in rigid spatial confinement
The extra-cellular microenvironment has a fundamental role in tumor growth and progression,
strongly affecting the migration strategies adopted by single cancer cells during metastatic invasion. In
this study, we use a novel microfluidic device to investigate the ability of mesenchymal and epithelial
breast tumor cells to fluidize and migrate through narrowing microstructures upon chemoattractant
stimulation. We compare the migration behavior of two mesenchymal breast cancer cell lines and one
epithelial cell line, and find that the epithelial cells are able to migrate through the narrowest
microconstrictions as the more invasive mesenchymal cells. In addition, we demonstrate that
migration of epithelial cells through a highly compressive environment can occur in absence of a
chemoattractive stimulus, thus evidencing that they are just as prone to react to mechanical cues as
invasive cell
Complex thermorheology of living cells
Temperature has a reliable and nearly instantaneous influence on mechanical responses of cells. As recently published, MCF-10A normal epithelial breast cells follow the time-temperature superposition (TTS) principle. Here, we measured thermorheological behaviour of eight common cell types within physiologically relevant temperatures and applied TTS to creep compliance curves. Our results showed that superposition is not universal and was seen in four of the eight investigated cell types. For the other cell types, transitions of thermorheological responses were observed at 36 °C. Activation energies (EA) were calculated for all cell types and ranged between 50 and 150 kJ mol-1. The scaling factors of the superposition of creep curves were used to group the cell lines into three categories. They were dependent on relaxation processes as well as structural composition of the cells in response to mechanical load and temperature increase. This study supports the view that temperature is a vital parameter for comparing cell rheological data and should be precisely controlled when designing experiments
Differences in cortical contractile properties between healthy epithelial and cancerous mesenchymal breast cells
Cell contractility is mainly imagined as a force dipole-like interaction based on actin stress fibers
that pull on cellular adhesion sites. Here, we present a different type of contractility based on
isotropic contractions within the actomyosin cortex. Measuring mechanosensitive cortical
contractility of suspended cells among various cell lines allowed us to exclude effects caused by
stress fibers. We found that epithelial cells display a higher cortical tension than mesenchymal cells,
directly contrasting to stress fiber-mediated contractility. These two types of contractility can even
be used to distinguish epithelial from mesenchymal cells. These findings from a single cell level
correlate to the rearrangement effects of actomyosin cortices within cells assembled in
multicellular aggregates. Epithelial cells form a collective contractile actin cortex surrounding
multicellular aggregates and further generate a high surface tension reminiscent of tissue
boundaries. Hence, we suggest this intercellular structure as to be crucial for epithelial tissue
integrity. In contrast, mesenchymal cells do not form collective actomyosin cortices reducing
multicellular cohesion and enabling cell escape from the aggregates
Mechano-Dependent Phosphorylation of the PDZ-Binding Motif of CD97/ADGRE5 Modulates Cellular Detachment
Summary Cells respond to mechanical stimuli with altered signaling networks. Here, we show that mechanical forces rapidly induce phosphorylation of CD97/ADGRE5 (pCD97) at its intracellular C-terminal PDZ-binding motif (PBM). Biochemically, this phosphorylation disrupts CD97 binding to PDZ domains of the scaffold protein DLG1. In shear-stressed cells, pCD97 appears not only in junctions, retracting fibers, and the attachment area but also in lost membrane patches, demonstrating (intra)cellular detachment at the CD97 PBM. This motif is critical for the CD97-dependent mechanoresponse. Cells expressing CD97 without the PBM are more deformable, and under shear stress, these cells lose cell contacts faster and show changes in the actin cytoskeleton when compared with cells expressing full-length CD97. Our data indicate CD97 linkage to the cytoskeleton. Consistently, CD97 knockout phenocopies CD97 without the PBM, and membranous CD97 is organized in an F-actin-dependent manner. In summary, CD97 shapes the cellular mechanoresponse through signaling modulation via its PBM
Complex thermorheology of living cells
Temperature has a reliable and nearly instantaneous influence on mechanical responses of cells. As recently published, MCF-10A normal epithelial breast cells follow the time-temperature superposition (TTS) principle. Here, we measured thermorheological behaviour of eight common cell types within physiologically relevant temperatures and applied TTS to creep compliance curves. Our results showed that superposition is not universal and was seen in four of the eight investigated cell types. For the other cell types, transitions of thermorheological responses were observed at 36 °C. Activation energies (EA) were calculated for all cell types and ranged between 50 and 150 kJ mol-1. The scaling factors of the superposition of creep curves were used to group the cell lines into three categories. They were dependent on relaxation processes as well as structural composition of the cells in response to mechanical load and temperature increase. This study supports the view that temperature is a vital parameter for comparing cell rheological data and should be precisely controlled when designing experiments
Actin and microtubule networks contribute differently to cell response for small and large strains
Cytoskeletal filaments provide cells with mechanical stability and organization. The main key players
are actin filaments and microtubules governing a cell’s response to mechanical stimuli. We
investigated the specific influences of these crucial components by deforming MCF-7 epithelial cells at
small(\u845% deformation) and large strains(>5% deformation). To understand specific contributions
of actin filaments and microtubules, we systematically studied cellular responses after treatment with
cytoskeleton influencing drugs. Quantification with the microfluidic optical stretcher allowed
capturing the relative deformation and relaxation of cells under different conditions. We separated
distinctive deformational and relaxational contributions to cell mechanics for actin and microtubule
networks for two orders of magnitude of drug dosages. Disrupting actin filaments via latrunculin A,
for instance, revealed a strain-independent softening. Stabilizing these filaments by treatment with
jasplakinolide yielded cell softening for small strains but showed no significant change at large strains.
In contrast, cells treated with nocodazole to disrupt microtubules displayed a softening at large strains
but remained unchanged at small strains. Stabilizing microtubules within the cells via paclitaxel
revealed no significant changes for deformations at small strains, but concentration-dependent
impact at large strains. This suggests that for suspended cells, the actin cortex is probed at small strains,
while at larger strains; the whole cell is probed with a significant contribution from the microtubule