Upon temperature changes, we observe a systematic shift of creep compliance curves J (t) for single living breast epithelial cells. We use a dual-beam laser trap (optical stretcher) to induce temperature jumps within milliseconds, while simultaneously measuring the mechanical response of whole cells to optical force. The cellular mechanical response was found to differ between sudden temperature changes compared to slow, long-term changes implying adaptation of cytoskeletal structure. Interpreting optically induced cell deformation as a thermorheological experiment allows us to consistently explain data on the basis of time-temperature superposition, well known from classical polymer physics. Measured time shift factors give access to the activation energy of the viscous flow of MCF-10A breast cells, which was determined to be ≈ 80 kJ mol −1 . The presented measurements highlight the fundamental role that temperature plays for the deformability of cellular matter. We propose thermorheology as a powerful concept to assess the inherent material properties of living cells and to investigate cell regulatory responses upon environmental changes.

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 (5% 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 microtubules.

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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 (E A ) 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.

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.

Solvent conditions are unexpectedly sufficient to drastically and reversibly slow down cells. In vitro on the molecular level, protein–solvent interactions drastically change in the presence of heavy water (D2O) and its stronger hydrogen bonds. Adding D2O to the cell medium of living cells increases the molecular intracellular viscosity. While cell morphology and phenotype remain unchanged, cellular dynamics transform into slow motion in a changeable manner. This is exemplified in the slowdown of cell proliferation and migration, which is caused by a reversible gelation of the cytoplasm. In analogy to the time–temperature superposition principle, where temperature is replaced by D2O, an increase in viscosity slows down the effective time. Actin networks, crucial structures in the cytoplasm, switch from a power‐law‐like viscoelastic to a more rubber‐like elastic behavior. The resulting intracellular resistance and dissipation impair cell movement. Since cells are highly adaptive non‐equilibrium systems, they usually respond irreversibly from a thermodynamic perspective. D2O induced changes, however, are fully reversible and their effects are independent of signaling as well as expression. The stronger hydrogen bonds lead to glass‐like, drawn‐out intramolecular dynamics, which may facilitate longer storage times of biological matter, for instance, during transport of organ transplants.

Oral squamous cell carcinomas (OSCC) are the 6 th most common cancer and the diagnosis is often belated for a curative treatment. The reliable and early differentiation between healthy and diseased cells is the main aim of this study in order to improve the quality of the treatment and to understand tumour pathogenesis.Here, the optical stretcher is used to analyse mechanical properties of cells and their potential to serve as a marker for malignancy. Stretching experiments revealed for the first time that cells of primary OSCCs were deformed by 2.9 % rendering them softer than cells of healthy mucosa which were deformed only by 1.9 %. Furthermore, the relaxation behaviour of the cells revealed that these malignant cells exhibit a faster contraction than their benign counterparts. This suggests that deformability as well as relaxation behaviour can be used as distinct parameters to evaluate emerging differences between these benign and malignant cells. Since many studies in cancer research are performed with cancer cell lines rather than primary cells, we have compared the deformability and relaxation of both types, showing that long time culturing leads to softening of cells. The higher degree of deformability and relaxation behaviour can enable cancer cells to traverse tissue emphasizing that changes in cell architecture may be a potential precondition for malignant transformation. Respecting the fact that even short culture times have an essential effect on the significance of the results, the use of primary cells for further research is recommended. The distinction between malignant and benign cells would enable an early confirmation of cancer diagnoses by testing cell samples of suspect oral lesions.