Application of sequential cyclic compression on cancer cells in a flexible microdevice

Abstract

Mechanical forces shape physiological structure and function within cell and tissue microenvironments, during which cells strive to restore their shape or develop an adaptive mechanism to maintain cell integrity depending on strength and type of the mechanical loading. While some cells are shown to experience permanent plastic deformation after a repetitive mechanical tensile loading and unloading, the impact of such repetitive compression on deformation of cells is yet to be understood. As such, the ability to apply cyclic compression is crucial for any experimental setup aimed at the study of mechanical compression taking place in cell and tissue microenvironments. Here, we demonstrate such cyclic compression using a microfluidic compression platform on live cell actin in SKOV-3 ovarian cancer cells. Live imaging of the actin cytoskeleton dynamics of the compressed cells was performed for varying pressures applied sequentially in ascending order during cell compression. Additionally, recovery of the compressed cells was investigated by capturing actin cytoskeleton and nuclei profiles of the cells at zero time and 24 h-recovery after compression in end point assays. This was performed for a range of mild pressures within the physiological range. Results showed that the phenotypical response of compressed cells during recovery after compression with 20.8 kPa differed observably from that for 15.6 kPa. This demonstrated the ability of the platform to aid in the capture of differences in cell behaviour as a result of being compressed at various pressures in physiologically relevant manner. Differences observed between compressed cells fixed at zero time or after 24 h-recovery suggest that SKOV-3 cells exhibit deformations at the time of the compression, a proposed mechanism cells use to prevent mechanical damage. Thus, biomechanical responses of SKOV-3 ovarian cancer cells to sequential cyclic compression and during recovery after compression could be revealed in a flexible microdevice. As demonstrated in this work, the observation of morphological, cytoskeletal and nuclear differences in compressed and non-compressed cells, with controlled micro-scale mechanical cell compression and recovery and using live-cell imaging, fluorescent tagging and end point assays, can give insights into the mechanics of cancer cells.

4D Force Detection of Cell Adhesion and Contractility

“…Therefore, the applied force of 1 μΝ resulted in an apical compression of ∼18 kPa. This value is in the range of physiological pressure reported for cancer cells . In the HLAC experiments, the cell area and traction force exerted by probed individual cells did not vary significantly before and after the compression cycles (SI Figure 4).…”

“…This value is in the range of physiological pressure reported for cancer cells. 45 In the HLAC experiments, the cell area and traction force exerted by probed individual cells did not vary significantly before and after the compression cycles (SI Figure 4). Additionally, the full viability of the HLAC protocol was demonstrated by a live/dead assay (SI Figure 5).…”

On-chip non-contact mechanical cell stimulation - quantification of SKOV-3 alignment to suspended microstructures

“…In addition to its dynamic compression capability, which was reported in our previous work [14, 15], the micro-piston device can be used simply in its static condition by employing the micro-piston as a hanging structure separated from the cells via a liquid-filled vertical gap. Typically, the device remains in this configuration during cell loading and mono-layer formation in preparation for compression studies.…”

“…, 250R and 280R, respectively. On the plot, n = 26,29, 39, 33, 42, 38,30,27, 33, 35, 36, 41,15, 36,21, 34, 48, 52 cells, respectively. Indicators on horizontal bars show significances of differences between cell elongation angle distribution in the groups (𝜒 2 test), *: p < 0.05, **: p < 0.003, ***: p < 0.0001, ns: not significant.…”

“…The frequencies of angle groups are shown in percentages over days (Day 0, 1, and 2) for different diameters of circular micro-piston of 200, 250, and 280 µm and their corresponding random/control area (R) of 200R, 250R and 280R, respectively. On the plot, n = 26,29, 39, 33, 42, 38,30,27, 33, 35, 36, 41,15, 36,21, 34, 48, 52 cells, respectively. Indicators on horizontal bars show significances of differences between cell elongation angle distribution in the groups (𝜒 2 test), *: p < 0.05, **: p < 0.003, ***: p < 0.0001, ns: not significant.…”