Jun 28, 2022, 2:50 PM
Room: S1

Room: S1


Micalet, Auxtine (University College London )


INTRODUCTION: Most epithelial cancer cell populations undergo an epithelial to mesenchymal transition (EMT), acquiring a more aggressive phenotype1. Mesenchymal cells are more motile and have the ability to remodel the extracellular matrix. This mechanical interaction with the surrounding matrix can be measured by bulk and single cell force generation. In this study we aim to use 3D in vitro methods to assess the contractility signatures of various epithelial cancer cell-lines.

Contraction assays. Two colorectal cancer (HT-29, HCT 116) and two breast cancer (MDA-MB-231, MCF-7) cell-lines were incorporated into collagen type I hydrogels at 1x106 cells/mL. Contraction was observed over 96 hours. Human Dermal Fibroblasts (HDFs) serve as a positive control.

Traction force microscopy (TFM). Polyacrylamide (PA) gels containing 0.1 μm red fluorescent beads were cast on glass bottom petri dishes. The gels were functionalised using collagen type I. Cells were seeded on the gels at a density of 1.5x105 cells and incubated for 24 hours before imaging. Comparative images were taken before and 15 minutes after trypsinization of the cells.

Quantitative polymerase chain reaction (qPCR). RNA was extracted from hydrogels through the TRI- Reagent® phase-separation method2. The following genes were measured for relative expression: LOX and RAE1.

Analysis. Images were analysed using Fiji ImageJ software and statistical analysis were performed using GraphPad Prism 9.

RESULTS: HDF cells contract the collagen type I hydrogels by 60% over 24 hours. MDA-MB-231 cells cause a 22% contraction whilst HT-29, HCT 116 and MCF-7 cells did not contract the gel. TFM results validate this trend showing that HDF cells displace the matrix by 2.8±1.7 µm. MDA-MB-231 cells displace the matrix by 0.7±0.1 µm. HCT 116, HT-29 and MCF-7 cells do not displace the matrix. MDA-MB-231 cells has significant upregulation of the EMT markers LOX and RAE1 compared to HT-29 cells (p=0.0070 and p=0.0237 respectively).

CONCLUSION: This study showed a correlation between the contractility profiles of epithelial cancer cells and their EMT status: highly mesenchymal-like cancer cells such as MDA-MB-231 cells are force-generating cells. This suggests that these cells can remodel the extracellular matrix, which aids migration and hence makes them highly invasive cancer cells.

ACKNOWLEDGEMENTS: MCF-7 cells were kindly provided by Dr Nina Moderau and Mr Michael Toeller from Imperial College London. We are grateful for supports from UCL Institute of Healthcare Engineering and EPSRC DTP PhD Studentship.

1. Hanahan, D. & Weinberg, R. A. Hallmarks of cancer: The next generation. Cell vol. 144 646–674 (2011).
2. DC, R., M, A., GJ, H. & TW, N. Purification of RNA using TRIzol (TRI reagent). Cold Spring Harb. Protoc. (2010).


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