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Multicellular architecture of malignant breast epithelia influences mechanics.

Venugopalan G, Camarillo DB, Webster KD, Reber CD, Sethian JA, Weaver VM, Fletcher DA, El-Samad H, Rycroft CH - PLoS ONE (2014)

Bottom Line: Using creep compression tests on multicellular breast epithelial structures, we found that pre-malignant acini with no lumen (MCF10AT) were significantly stiffer than normal hollow acini (MCF10A) by 60%.This difference depended on structural changes in the pre-malignant acini, as neither single cells nor normal multicellular acini tested before lumen formation exhibited these differences.We also simulated a single contracting cell in different multicellular architectures and found that lumen filling led to a 20% increase in the "perceived stiffness" of a single contracting cell independent of any changes to matrix mechanics.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering and Biophysics Program, University of California, Berkeley, California, United States of America.

ABSTRACT
Cell-matrix and cell-cell mechanosensing are important in many cellular processes, particularly for epithelial cells. A crucial question, which remains unexplored, is how the mechanical microenvironment is altered as a result of changes to multicellular tissue structure during cancer progression. In this study, we investigated the influence of the multicellular tissue architecture on mechanical properties of the epithelial component of the mammary acinus. Using creep compression tests on multicellular breast epithelial structures, we found that pre-malignant acini with no lumen (MCF10AT) were significantly stiffer than normal hollow acini (MCF10A) by 60%. This difference depended on structural changes in the pre-malignant acini, as neither single cells nor normal multicellular acini tested before lumen formation exhibited these differences. To understand these differences, we simulated the deformation of the acini with different multicellular architectures and calculated their mechanical properties; our results suggest that lumen filling alone can explain the experimentally observed stiffness increase. We also simulated a single contracting cell in different multicellular architectures and found that lumen filling led to a 20% increase in the "perceived stiffness" of a single contracting cell independent of any changes to matrix mechanics. Our results suggest that lumen filling in carcinogenesis alters the mechanical microenvironment in multicellular epithelial structures, a phenotype that may cause downstream disruptions to mechanosensing.

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Single cell mechanics and cell–cell connections do not explain the mechanical differences.Creep compliance (mean ±95% CI) of MCF10A and MCF10AT cells at (A) single cell state (N = 14 and N = 15 cells for A and T respectively) and (B) 6–8 day state before lumen formation (N = 34 and N = 33 colonies for A and T respectively). Confocal immunofluorescence images of 8 day colonies of (C) MCF10A and (D) MCF10AT; 6–8 day time points were selected for testing because this was before lumina formed; scale bars 25 µm.
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pone-0101955-g003: Single cell mechanics and cell–cell connections do not explain the mechanical differences.Creep compliance (mean ±95% CI) of MCF10A and MCF10AT cells at (A) single cell state (N = 14 and N = 15 cells for A and T respectively) and (B) 6–8 day state before lumen formation (N = 34 and N = 33 colonies for A and T respectively). Confocal immunofluorescence images of 8 day colonies of (C) MCF10A and (D) MCF10AT; 6–8 day time points were selected for testing because this was before lumina formed; scale bars 25 µm.

Mentions: To test the potential contribution of mechanical changes at the single cell level, we embedded MCF10A and MCF10AT cells in laminin-rich ECM as before, but extracted them after twelve hours and subjected single cells to creep compression tests. MCF10AT single cells were not noticeably stiffer than MCF10A single cells (one-sided t-test, p = 0.329), suggesting that the increased stiffness observed for pre-malignant acini does not result from stiffer cells (Fig. 3A). To determine the influence of cell–cell connectivity strength, we extracted MCF10A and MCF10AT structures after 6–8 days of growth. As suggested by previous literature [20], 6–8 day-old MCF10A structures did not yet have lumina (i.e. acini were filled, Figure 3C,D). At this time point, healthy and pre-malignant structures had the same filled multicellular architecture, and did not exhibit distinguishable differences in creep compliance (Fig. 3B, p = 0.963). Since changes in cell–cell connectivity would be present at the 6–8 day time point, these data suggest that neither single cell mechanics nor cell–cell connectivity can account for the decreased compliance observed in pre-malignant structures. Notably, both of the 6–8 day filled structures (Fig. 3B) exhibited similar creep responses to mature MCF10AT acini (Fig. 2C). Assuming that cell–cell connectivity remains similar in the 6–8 day acini and the mature acini, the data suggests that the decreased stiffness of the acini coincides with hollow lumen formation.


Multicellular architecture of malignant breast epithelia influences mechanics.

Venugopalan G, Camarillo DB, Webster KD, Reber CD, Sethian JA, Weaver VM, Fletcher DA, El-Samad H, Rycroft CH - PLoS ONE (2014)

Single cell mechanics and cell–cell connections do not explain the mechanical differences.Creep compliance (mean ±95% CI) of MCF10A and MCF10AT cells at (A) single cell state (N = 14 and N = 15 cells for A and T respectively) and (B) 6–8 day state before lumen formation (N = 34 and N = 33 colonies for A and T respectively). Confocal immunofluorescence images of 8 day colonies of (C) MCF10A and (D) MCF10AT; 6–8 day time points were selected for testing because this was before lumina formed; scale bars 25 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4128597&req=5

pone-0101955-g003: Single cell mechanics and cell–cell connections do not explain the mechanical differences.Creep compliance (mean ±95% CI) of MCF10A and MCF10AT cells at (A) single cell state (N = 14 and N = 15 cells for A and T respectively) and (B) 6–8 day state before lumen formation (N = 34 and N = 33 colonies for A and T respectively). Confocal immunofluorescence images of 8 day colonies of (C) MCF10A and (D) MCF10AT; 6–8 day time points were selected for testing because this was before lumina formed; scale bars 25 µm.
Mentions: To test the potential contribution of mechanical changes at the single cell level, we embedded MCF10A and MCF10AT cells in laminin-rich ECM as before, but extracted them after twelve hours and subjected single cells to creep compression tests. MCF10AT single cells were not noticeably stiffer than MCF10A single cells (one-sided t-test, p = 0.329), suggesting that the increased stiffness observed for pre-malignant acini does not result from stiffer cells (Fig. 3A). To determine the influence of cell–cell connectivity strength, we extracted MCF10A and MCF10AT structures after 6–8 days of growth. As suggested by previous literature [20], 6–8 day-old MCF10A structures did not yet have lumina (i.e. acini were filled, Figure 3C,D). At this time point, healthy and pre-malignant structures had the same filled multicellular architecture, and did not exhibit distinguishable differences in creep compliance (Fig. 3B, p = 0.963). Since changes in cell–cell connectivity would be present at the 6–8 day time point, these data suggest that neither single cell mechanics nor cell–cell connectivity can account for the decreased compliance observed in pre-malignant structures. Notably, both of the 6–8 day filled structures (Fig. 3B) exhibited similar creep responses to mature MCF10AT acini (Fig. 2C). Assuming that cell–cell connectivity remains similar in the 6–8 day acini and the mature acini, the data suggests that the decreased stiffness of the acini coincides with hollow lumen formation.

Bottom Line: Using creep compression tests on multicellular breast epithelial structures, we found that pre-malignant acini with no lumen (MCF10AT) were significantly stiffer than normal hollow acini (MCF10A) by 60%.This difference depended on structural changes in the pre-malignant acini, as neither single cells nor normal multicellular acini tested before lumen formation exhibited these differences.We also simulated a single contracting cell in different multicellular architectures and found that lumen filling led to a 20% increase in the "perceived stiffness" of a single contracting cell independent of any changes to matrix mechanics.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering and Biophysics Program, University of California, Berkeley, California, United States of America.

ABSTRACT
Cell-matrix and cell-cell mechanosensing are important in many cellular processes, particularly for epithelial cells. A crucial question, which remains unexplored, is how the mechanical microenvironment is altered as a result of changes to multicellular tissue structure during cancer progression. In this study, we investigated the influence of the multicellular tissue architecture on mechanical properties of the epithelial component of the mammary acinus. Using creep compression tests on multicellular breast epithelial structures, we found that pre-malignant acini with no lumen (MCF10AT) were significantly stiffer than normal hollow acini (MCF10A) by 60%. This difference depended on structural changes in the pre-malignant acini, as neither single cells nor normal multicellular acini tested before lumen formation exhibited these differences. To understand these differences, we simulated the deformation of the acini with different multicellular architectures and calculated their mechanical properties; our results suggest that lumen filling alone can explain the experimentally observed stiffness increase. We also simulated a single contracting cell in different multicellular architectures and found that lumen filling led to a 20% increase in the "perceived stiffness" of a single contracting cell independent of any changes to matrix mechanics. Our results suggest that lumen filling in carcinogenesis alters the mechanical microenvironment in multicellular epithelial structures, a phenotype that may cause downstream disruptions to mechanosensing.

Show MeSH
Related in: MedlinePlus