Trade-off movement between hydraulic resistance escape and shear stress escape by cancer cells

Abstract

In the circulatory system, the microenvironment surrounding cancer cells is complex and involves multiple coupled factors. We selected two core physical factors, shear stress and hydraulic resistance, and constructed a microfluidic device with dual negative inputs to study the trade-off movement behavior of cancer cells when facing coupled factors. We detected significant shear stress escape phenomena in the MDA-MB-231 cell line and qualitatively explained this behavior using a cellular force model. Through the dual validation of substrate anti-cell-adhesion modification and employment of the MCF-7 cell line, we further substantiated the predictability and feasibility of our model. This study provides an explanation for the trade-off underlying the direction-choosing mechanism of cancer cells when facing environmental selection.

Figure 1

Specific structure of the microfluidic device and verification of interstitial flow. (A) The structure of the functional microfluidic chip. (B) Schematic diagram of the fabrication process. (C) Schematic of the fluorescence image and the extracted fluorescence intensity when the channel was filled with dextran-FITC and the cell was at the branch. (D) Statistical analysis of the fluorescence intensity of the background, the channel containing cells and the channel without cells, and the leakage ratio of the channel with n > 20 cells. Scale bars, 50 μm.

Figure 2

Decision-making characteristics of the cells at branches. (A) Normalized area ratio and final direction selection of the cells at 0.5 (top) and 3 mbar/cell (bottom). (B) Schematic diagram of cell migration within the channel. Pressure condition: 0.5 mbar/cell. (i) Representative cell entering the inlet of the pipette. (ii) Representative cell at the branch and making trade-off choices. (iii) Representative cell entering 4×. (iv) Representative image of a cell entering 1×. (C) Decision-making time of the cells at 0.5 (left) and 3 mbar/cell (right). Red: cells entering the 4× channel; gray: cells entering the 1× channel. The data are presented as the means ± SDs, and statistical comparisons were made via Student’s t-test; ^∗p < 0.05 and n.s. p > 0.05. (D) Inlet-to-branch time of the cells at different pressures. Inset: fit function and correlation coefficient. (E) Decision-making time of the cells at different pressures. The number of cells was n > 90. Scale bars, 50 μm.

Figure 3

Trade-offs between the escape and motility of MDA-MB-231 cells. (A) Percentages of cells that entered the 4× channel and 1× channel at different pressures in the vehicle control (VC) and cells that responded to blebbistatin (repeat experiments n ≥ 3). (B) Representative cells exhibiting the protrusion dynamics of blebs and protrusions. Pressure condition: 0.5 mbar/cell. (C) Statistical analysis of the transition mode proportions into different channels under different pressures (repeat experiments n = 3). (D) Representative image sequence depicting the Rac1-YFP–labeled MDA-MB-231 cells in each branch at 0.5 mbar/cell. Yellow arrows: fluorescence enrichment in the channel selected. (E) Normalized fluorescence intensity at 0.5 and 3 mbar/cell (cell numbers n > 40). The data represent the means ± SDs, with statistical comparisons performed via Student’s t-test; ^∗∗∗∗p < 0.0001 and n.s. p > 0.05. Scale bars, 50 μm.

Figure 4

Schematic diagram and simulation of the forces during trade-off escape. (A) Force labeling of cancer cells on both sides of the channels. (B) Sketch map of the trade-off between hydraulic resistance and shear stress. (C) Simulation of the variation in the shear-stress-induced active force and passive force with pressure under the given parameters. (D) Force difference and coupling force between the 4× and 1× channels.

Figure 5

Force numerical simulation and experimental validation for different F_max values. (A and B) Numerical simulation of the shear-stress-induced active force and coupling force for different F_max and κ_R values. (C and D) Inlet-to-branch time and decision-making time of the MCF-7 cell line and MDA-MB-231 cell line for the initial 20 h of PLL-g-PEG substrate treatment (n > 40 cells). Inset: fit function and correlation coefficient. (E) Percentages of the cells that entered the 4× channel and 1× channel at different pressures for the MCF-7 cell line (repeat experiments n = 3). (F) Percentages of cells that entered the 4× channel and 1× channel during the initial 20 h under PLL-g-PEG substrate treatment (repeat experiments n = 3).