Transient external force induces phenotypic reversion of malignant epithelial structures via nitric oxide signaling
- University of California, Berkeley, United States
- Lawrence Berkeley National Laboratory, United States
- National Cancer Institute, National Institutes of Health, United States
Figures
Figure 1 with 4 supplements
Transient compression of breast epithelial cells grown in laminin-rich ECM reverts the malignant phenotype.
(A) Cells were compressed after 30 min of lrECM polymerization at 37°C, before first division (day 1) or formation of growth-arrested colonies or large disorganized colonies (day 10). Compressive …
-
Figure 1—source data 1
Colony size and proliferation status in compressed and uncompressed T4-2 and S1 cells.
- https://doi.org/10.7554/eLife.26161.007
Figure 1—figure supplement 1
Application of transient compression to 3D cultured breast epithelial cells.
(F) Photograph of PDMS chamber for applying compression to 3D cultured cells, demonstrating the unstretched and pre-stretched configurations. In an experiment, one well was pre-stretched and held in …
Figure 1—figure supplement 2
Mechanical reversion by transient compression occurs in both 3D embedded and 3D-on-top culture geometries.
(I) 23% compression of single malignant cells led to reduction in colony size. *Difference is significant at p<0.05 in one-way analysis of variance with Tukey-Kramer multiple comparison test. Each …
Figure 1—figure supplement 3
Chemical agents revert the malignant phenotype.
Representative confocal immunofluorescence images of T4-2 colonies after 10 days of embedded culture in lrECM with (K) no drug treatment, (L) the EGFR inhibitor Tyrphostin AG 1478 (70 nM) or (M) …
Figure 1—figure supplement 4
Transient compression reduces growth of in cultured malignant cells of the MCF10A series.
(N) Brightfield images of compressed and uncompressed cells of the MCF10A breast cancer progression series. Non-malignant MCF10A cells and malignant MCF10A-CA1d cells were seeded and compressed in …
Figure 2
Mechanical reversion occurs above a threshold strain.
(A) Bright-field and confocal immunofluorescence images of malignant T4-2 cell colonies grown for 10 days in compressed gels. Scale bars 100 μm bright-field and 25 μm confocal. (B) Colonies grown in …
-
Figure 2—source data 1
T4-2 colony size and architecture by compressive strain.
- https://doi.org/10.7554/eLife.26161.009
Figure 3 with 1 supplement
Mechanical reversion occurs via nitric oxide signaling.
(A) Malignant T4-2 cells were compressed 30 min after seeding and coating with 5% lrECM drip in the presence of the nitric oxide (NO) sensor dye DAF-FM DA. At the indicated times after compression, …
-
Figure 3—source data 1
Nitric oxide sensor in T4-2 cells and nitric oxide synthase inhibitor-treated T4-2 colonites.
- https://doi.org/10.7554/eLife.26161.012
Figure 3—figure supplement 1
Pairwise comparison of colony sizes while manipulating nitric oxide conditions and mechanical stimulation.
Cross-sectional area of T4-2 colonies after 5 days of culture under a given pair of condition across individual experiments. Each point is the mean colony size of 159 ± 42 colonies (mean ± SD; range …
Figure 4 with 3 supplements
Compression allows restoration of coherent rotation after division.
(A) Time series of malignant T4-2 cells growing in uncompressed and compressed gels. Scale bar 10 μm. Time series were taken from Figure 4—Video 1 and Figure 4—Video 1. (B) Cells at the two-cell …
-
Figure 4—source data 1
- https://doi.org/10.7554/eLife.26161.015
Figure 4—figure supplement 1
E-cadherin function is required for mechanical reversion.
Single malignant T4-2 cells were treated with function-blocking E-cadherin antibody before compression was applied (E) Bright-field images of colonies treated with function-blocking E-cadherin …
Figure 4—video 1
Uncompressed malignant T4-2 doublets undergo incoherent movement.
Time-lapse of uncompressed malignant T4-2 breast epithelial cells. Scale bar 25 μm, time stamp hours:minutes.
Figure 4—video 2
Compression restores coherent rotation of malignant T4-2 doublets.
Time-lapse of compressed malignant T4-2 breast epithelial cells. Scale bar 25 μm, time stamp hours:minutes.
Additional files
-
Transparent reporting form
- https://doi.org/10.7554/eLife.26161.018
