Compressive stress triggers fibroblasts spreading over cancer cells to generate carcinoma in situ organization.

At the early stage of tumor progression, fibroblasts are located at the outer edges of the tumor, forming an encasing layer around it. In this work, we have developed a 3D in vitro model where fibroblasts' layout resembles the structure seen in carcinoma in situ. We use a microfluidic encapsulation technology to co-culture fibroblasts and cancer cells within hollow, permeable, and elastic alginate shells. We find that in the absence of spatial constraint, fibroblasts and cancer cells do not mix but segregate into distinct aggregates composed of individual cell types. However, upon confinement, fibroblasts enwrap cancer cell spheroid. Using a combination of biophysical methods and live imaging, we find that buildup of compressive stress is required to induce fibroblasts spreading over the aggregates of tumor cells. We propose that compressive stress generated by the tumor growth might be a mechanism that prompts fibroblasts to form a capsule around the tumor.

Mechanical forces across compartments coordinate cell shape and fate transitions to generate tissue architecture.

Morphogenesis and cell state transitions must be coordinated in time and space to produce a functional tissue. An excellent paradigm to understand the coupling of these processes is mammalian hair follicle development, which is initiated by the formation of an epithelial invagination-termed placode-that coincides with the emergence of a designated hair follicle stem cell population. The mechanisms directing the deformation of the epithelium, cell state transitions and physical compartmentalization of the placode are unknown. Here we identify a key role for coordinated mechanical forces stemming from contractile, proliferative and proteolytic activities across the epithelial and mesenchymal compartments in generating the placode structure. A ring of fibroblast cells gradually wraps around the placode cells to generate centripetal contractile forces, which, in collaboration with polarized epithelial myosin activity, promote elongation and local tissue thickening. These mechanical stresses further enhance compartmentalization of Sox9 expression to promote stem cell positioning. Subsequently, proteolytic remodelling locally softens the basement membrane to facilitate a release of pressure on the placode, enabling localized cell divisions, tissue fluidification and epithelial invagination into the underlying mesenchyme. Together, our experiments and modelling identify dynamic cell shape transformations and tissue-scale mechanical cooperation as key factors for orchestrating organ formation.

SnapShot: Mechanotransduction in the nucleus.

Cells are continuously exposed to tissue-specific extrinsic forces that are counteracted by cell-intrinsic force generation through the actomyosin cytoskeleton and alterations in the material properties of various cellular components, including the nucleus. Forces impact nuclei both directly through inducing deformation, which is sensed by various mechanosensitive components of the nucleus, as well as indirectly through the actomyosin cytoskeleton and mechanosensitive pathways activated in the cytoplasm. To view this SnapShot, open or download the PDF.

Microfluidic-Based Generation of 3D Collagen Spheres to Investigate Multicellular Spheroid Invasion.

During tumor progression, cancer cells acquire the ability to escape the primary tumor and invade adjacent tissues. They migrate through the stroma to reach blood or lymphatics vessels that will allow them to disseminate throughout the body and form metastasis at distant organs. To assay invasion capacity of cells in vitro, multicellular spheroids of cancer cells, mimicking primary tumor, are commonly embedded in collagen I extracellular matrix, which mimics the stroma. However, due to their higher density, spheroids tend to sink at the bottom of the collagen droplets, resulting in the spreading of the cells on two dimensions. We developed an innovative method based on droplet microfluidics to embed and control the position of multicellular spheroids inside spherical droplets of collagen. In this method cancer cells are exposed to a uniform three-dimensional (3D) collagen environment resulting in 3D cell invasion.

Intracellular dynamics measurements with full field optical coherence tomography suggest hindering effect of actomyosin contractility on organelle transport.

Intracellular motion can be quantitatively monitored in tissues using coherence-gated microscopic techniques. With full-field optical coherence tomography (FFOCT), the use of high numerical aperture microscope objectives provides a high resolution mapping of intracellular dynamics that are probed with subwavelength sensitivity. In the upper temporal bandwidth that we have used (1-6 Hz) the main contribution to the dynamic signal arises from the overall dynamical, optically heterogeneous cytoplasm. We propose a method to specifically study the impact of actomyosin contractility on the intracellular dynamic signal by performing high throughput, comparative measurements of multicellular aggregates with and without blebbistatin action, a selective inhibitor of class-II myosins that disrupts actomyosin contractile activity. Our results indicate a significant increase in the fraction of the signal that decorrelates within 1 second after inhibition of contractility. This observation mitigates the anticipated importance of actomyosin contractile forces to directly move organelles, but highlights their role in hindering organelle transport via their stiffening effect of the viscoelastic cytoplasm.