Abstract
Atomic Force Microscopy (AFM) is successfully used for the quantitative investigation of the cellular mechanosensing of the microenvironment. To this purpose, several force spectroscopy approaches aim at measuring the adhesive forces between two living cells and also between a cell and a suitable reproduction of the extracellular matrix (ECM), typically exploiting tips suitably functionalised with single components (e.g. collagen, fibronectin) of the ECM. However, these probes only poorly reproduce the complexity of the native cellular microenvironment and consequently of the biological interactions.
We developed a novel approach to produce AFM probes that faithfully reproduce the structural and biochemical complexity of the ECM; this was achieved by attaching to an AFM cantilever a micrometric slice of native decellularised ECM cut by laser microdissection, and preserving the full morphological, mechanical, and chemical heterogeneity of the ECM.
These native ECM probes can be used in force spectroscopy experiments aimed at targeting cell-microenvironment interactions. Here, we demonstrate the feasibility of dissecting mechanotransductive cell-ECM interactions in the 10 pN range. As proof-of-principle, we tested a rat bladder ECM probe against the AY-27 rat bladder cancer cell line. On the one hand, we obtained reproducible results using different probes derived from the same ECM regions; on the other hand, we detected differences in the adhesion patterns of distinct bladder ECM regions, such as submucosa and detrusor, in line with the disparities in composition and biophysical properties of these ECM regions.
Our results demonstrate that it is possible to use native ECM probes, produced from patient-specific regions of organs and tissues, to investigate cell-microenvironment interactions and early mechanotransductive processes by force spectroscopy. This opens new possibilities in the field of personalised medicine.
Competing Interest Statement
The authors have declared no competing interest.
