The cyto-linker and scaffolding protein "plectin" mis-localization leads to softening of cancer cells.

Discovering tools to prevent cancer progression requires understanding the fundamental differences between normal and cancer cells. More than a decade ago, atomic force microscopy (AFM) revealed cancer cells' softer body compared to their healthy counterparts. Here, we investigated the mechanism underlying the softening of cancerous cells in comparison with their healthy counterparts based on AFM high resolution stiffness tomography and 3D confocal microscopy. We showed microtubules (MTs) network in invasive ductal carcinoma cell cytoskeleton is basally located and segmented for around 400 nm from the cell periphery. Additionally, the cytoskeleton scaffolding protein plectin exhibits a mis-localization from the cytoplasm to the surface of cells in the carcinoma which justifies the dissociation of the MT network from the cell's cortex. Furthermore, the assessment of MTs' persistence length using a worm-like-chain (WLC) model in high resolution AFM images showed lower persistence length of the single MTs in ductal carcinoma compared to that in the normal state. Overall, these tuned mechanics support the invasive cells to ascertain more flexibility under compressive forces in small deformations. These data provide new insights into the structural origins of cancer aids in progression.

Probing local lateral forces of focal adhesions and cell-cell junctions of living cells by torsional force spectroscopy.

The number and strength of mechanical connections of cells to their local environment can be indicative of their migration and invasion potential. Gaining direct access to the mechanical properties of individual connections and bringing them into a relationship with the state of disease, however, is a formidable task. Here, we present a method to directly sense focal adhesions and cell-cell contacts with a force sensor to quantify the lateral forces of their anchoring points. We found local lateral forces of 1.0-1.5 nN for focal adhesions and slightly higher values at the interfaces between cells where cell-cell contacts are located. Interestingly, a modified surface layer was observed exhibiting considerably reduced tip friction directly next to the area of a retracting cell edge on the substrate. We expect that this technique can improve the understanding of the relationship between mechanical properties of cell connections and the pathological state of cells in the future.

Carcinomas with Occult Metastasis Potential: Diagnosis/Prognosis Accuracy Improvement by Means of Force Spectroscopy.

Accurate diagnosis of cancer stage is inevitable for the following prognosis in patients struggling with these lesions to promote their health and survival rate. Previous studies on survival rate statistics show, in some cases, failure in cancer stage surveys in which metastasis or recurrence of the disease was not accurately prognosed. Morphology study of cancer cells advances the understanding about cancer behavior and its progression, in which, in our previous study on invasive cancer cells, fewer formations of cytoskeleton components compared to their counterparts was observed. Here it is shown that carcinomas with an occult propensity of metastasis depict a number of poorly differentiated cells with decreased amounts of cytoskeleton components in a near-well differentiated population. Force spectroscopy in conjunction with fluorescence microscopy of lung cancer, liver hepatoma, and melanoma provides a general view of these cells' architecture, leading to the conclusion that the scarce abnormal-shaped cells with low formation of structural filaments convey the high risk of metastatic potential of the tumor. The results demonstrate that force spectroscopy complements conventional diagnostic approaches by an accurate cytoskeleton assessment and can improve the following prognosis in epithelial cancers with occult metastasis risk.

Structural analysis of healthy and cancerous epithelial-type breast cells by nanomechanical spectroscopy allows us to obtain peculiarities of the skeleton and junctions.

The transition of healthy epithelial cells to carcinoma is associated with an alteration in the structure and organization of the cytoskeleton of the cells. A comparison of the mechanical properties of cancerous and healthy cells indicated a higher deformability of the cancer cells based on averaging the mechanical properties of single cells. However, the exact reason for softening of the cancerous cells compared to their counterparts remains unclear. Here, we focused on nanomechanical spectroscopy of healthy and cancerous ductal epithelial-type breast cells by means of atomic force microscopy with high lateral and depth precision. As a result, based on atomic force microscopy measurements formation of significantly fewer microtubules in cancerous cells which was observed in our study is most likely one of the main causes for the overall change in mechanical properties without any phenotypic shift. Strikingly, in a confluent layer of invasive ductal carcinoma cells, we observed the formation of cell-cell junctions that have the potential for signal transduction among neighboring cells such as desmosomes and adherens junctions. This increases the possibility of cancerous cell collaboration in malignancy, infiltration or metastasis phenomena.