This paper demonstrates a protocol to characterize the mechanical properties of living cells by means of microindentation using an...

Adult stem cells are essential for tissue maintenance and repair. Although genetic pathways for controlling adult stem cells are extensively investigated in various tissues, much less is known about how mechanosensing could regulate adult stem cells and tissue growth. Here, we demonstrate that shear stress sensing regulates intestine stem cell proliferation and epithelial cell number in adult Drosophila . Ca ²⁺ imaging in ex vivo midguts shows that shear stress, but not other mechanical forces, specifically activates enteroendocrine cells among all epithelial cell types. This activation is mediated by transient receptor potential A1 (TrpA1), a Ca ²⁺ -permeable channel expressed in enteroendocrine cells. Furthermore, specific disruption of shear stress, but not chemical, sensitivity of TrpA1 markedly reduces proliferation of intestinal stem cells and midgut cell number. Therefore, we propose that shear stress may act as a natural mechanical stimulation to activate TrpA1 in enteroendocrine cells, which, in turn, regulates intestine stem cell behavior.

It has previously been shown that the simultaneous activation of PI3K (phosphatidylinositol 3-kinase) and Ras/MAPK (mitogen-activated protein kinases) pathways facilitate tumor growth despite only inducing cancer cell dormancy individually. Determining the impacts on cellular mechanics each pathway incites alone and in unison is critical to developing non-toxic cancer therapies for triple-negative breast cancers. PTEN (phosphatase and tensin homolog) knockout and activated KRAS (Kristen rat sarcoma viral oncogene homolog) overexpression in healthy MCF-10A human breast epithelial cells activated the PI3K and Ras/MAPK pathways, respectively. Cell stiffness and fluidity were simultaneously measured using atomic force microscopy. Results suggest that PTEN knockout reduced cell stiffness and increased cell fluidity independent of PI3K activation. Effects of activated KRAS overexpression on cell stiffness depends on rigidity of cell culture substrate. Activated KRAS overexpression also counteracts the effects of PTEN knockout.

Cancer cell migration through tissue pores and tracks into the bloodstream is a critical biological step for cancer metastasis. Although in vivo studies have shown that expression of vimentin can induce invasive cell lines, its role in cell cytoskeleton reorganization and cell motility under in vitro physical confinement remains unknown. Here, a microfluidic device with cell culture chamber and collagen-coated microchannels was developed as an in vitro model for physiological confinement environments. Using this microchannel assay, we demonstrated that the knockdown of vimentin decreases 3T3 fibroblast cell directional migration speed in confined microchannels. Additionally, as cells form dynamic membranes that define the leading edge of motile cells, different leading edge morphologies of 3T3 fibroblast and 3T3 vimentin knockdown cells were observed. The leading edge morphology change under confinement can be explained by the effect of vimentin on cytoskeletal organization and focal adhesion. The microfluidic device integrated with a time-lapse microscope provided a new approach to study the effect of vimentin on cell adhesion, migration, and invasiveness.