Mechanosensitive structures in epithelial integrity

Force-producing actomyosin bundles are required for the maintenance of intracellular tension at cell-cell junctions and cell-substrate interface of immotile epithelial sheets. Through their connections to both integrin-and cadherin-based cell adhesion sites, these mechanosensitive structures also sense changes in their physical environment and adjust cellular contractility in order to maintain a force-equilibrium with the surroundings. During neoplastic progression, abnormal regulation of these mechanosensitive structures may cause a force-imbalance between the cell and its environment, possibly creating favourable conditions for the transformed cells to escape from the primary tumor site. In our research, we are focusing on the mechanosensitive signaling pathways that regulate the assembly and maintenance of actomyosin structures and trying to find out how alterations in these pathways could result in scattering of the transformed epithelial cells. Additionally, we are trying to understand how the detached cancer cells utilize these aberrantly regulated actin-based structures in intruding to the surrounding tissues. In these studies we utilize several human and canine cell lines and manipulation of 2D and 3D cultures combined with advanced imaging methods.

In the image: MDCK monolayer, Actin-blue, P-MLC-green, DAPI-red

 

In­ter­play with stromal cells

Nowadays it is understood that cancer cells do not invade without the help of their stroma. Stromal cells undergo significant changes in their gene-expression profiles along cancer progression, signal abnormally to epithelial cells and may modify the micro-environment more permissive to cancer cell invasion. This abnormal stromal impact may induce invasive cellular features in transformed cells through their actomyosin machinery. We are currently investigating the role of stromal adipocytes in this process. These cell are the most abundant cell type in mammary gland stroma but still the least studied, despite the clear connection between excessive adipose and invasive breast cancer.

 

In the Image: Primary carcinosarcoma sample from the mammary gland of canine, containing epithelial cell population and fibroblasts

 

Bio­phys­ical changes of the tu­mor stroma

In most solid tumors cancer cells are exposed to major mechanical changes of the stroma. An overall stiffening of the surroundings may activate specific mechanosensitive pathways with subsequent alterations in the actomyosin bundles. Increasing stiffness is known to strongly correlate with the development of invasive breast cancer, suggesting that changes in the biophysical environment could promote invasive disease through the generation of abnormal cellular forces. We aim in understanding how changes in the biophysical properties of cell microenvironment affects the regulation of force-producing actomyosin bundles and how it is linked to the invasion potential of the cancer cells. In these studies we utilize both human and canine-derived cell lines in shape-and stiffness-determined 3D culture setups.

 

 

In the Image: Mammary spheroids formed by breast epithelial cells in compliant matrix

My­oep­ithelial cells and their role in can­cer in­va­sion

In majority of the breast cancers early phase of the disease involves neoplastic proliferation of the ductal epithelium, slowly progressing into ductal carcinoma in situ (DCIS). These events precede transition into more aggressive stage whereupon transformed cells adopt invasive features, overcoming the barrier made by myoepithelial (ME) cells and basement membrane. Myoepithelial cells have been suggested to protect against spreading of cancer cells at the early stages of the disease, although the mechanisms are not well understood. Our studies aim in revealing the molecular mechanisms behind the alterations of ME layer that are connected to cancer invasion. In these studies we utilize human primary myoepithelial cells as well as canine patient material from different stages of mammary carcinomas.

In the image: Smooth muscle actin (SMA) staining of a paraffin-embedded normal canine mammary gland tissue slice