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Abstract Peritoneal metastases (PM) are common routes of dissemination for colorectal cancer (CRC) and remain a lethal disease with a poor prognosis. The properties of the extracellular matrix (ECM) are important in cancer development; studying their changes is crucial to understand CRC-PM development. We studied the elastic properties of ECMs derived from human samples of normal and neoplastic PM by atomic force microscopy (AFM); results were correlated with patient clinical data and expression of ECM components related to metastatic spread. We show that PM progression is accompanied by stiffening of the ECM, increased cancer associated fibroblasts (CAF) activity and increased deposition and crosslinking in neoplastic matrices; on the other hand, softer regions are also found in neoplastic ECMs on the same scales. Our results support the hypothesis that local changes in the normal ECM can create the ground for growth and spread from the tumour of invading metastatic cells. We have found correlations between the mechanical properties (relative stiffening between normal and neoplastic ECM) of the ECM and patients’ clinical data, like age, sex, presence of protein activating mutations in BRAF and KRAS genes and tumour grade. Our findings suggest that the mechanical phenotyping of PM-ECM has the potential to predict tumour development.

Abstract The cell/microenvironment interface is the starting point of integrin-mediated mechanotransduction, but many details of mechanotransductive signal integration remain elusive due to the complexity of the involved (extra)cellular structures, such as the glycocalyx. We used nano-bio-interfaces reproducing the complex nanotopographical features of the extracellular matrix to analyse the glycocalyx impact on PC12 cell mechanosensing at the nanoscale (e.g., by force spectroscopy with functionalised probes). Our data demonstrates that the glycocalyx configuration affects spatio-temporal nanotopography-sensitive mechanotransductive events at the cell/microenvironment interface. Opposing effects of major glycocalyx removal were observed, when comparing flat and specific nanotopographical conditions. The excessive retrograde actin flow speed and force loading are strongly reduced on certain nanotopographies upon strong reduction of the native glycocalyx, while on the flat substrate we observe the opposite trend. Our results highlight the importance of the glycocalyx configuration in a molecular clutch force loading-dependent cellular mechanism for mechanosensing of microenvironmental nanotopographical features. Graphical Abstract

Biosensors are aimed at detecting tiny physical and chemical stimuli in biological systems. Physical forces are ubiquitous, being implied in all cellular processes, including cell adhesion, migration, and differentiation. Given the strong interplay between cells and their microenvironment, the extracellular matrix (ECM) and the structural and mechanical properties of the ECM play an important role in the transmission of external stimuli to single cells within the tissue. Vice versa, cells themselves also use self-generated forces to probe the biophysical properties of the ECM. ECM mechanics influence cell fate, regulate tissue development, and show peculiar features in health and disease conditions of living organisms. Force sensing in biological systems is therefore crucial to dissecting and understanding complex biological processes, such as mechanotransduction. Atomic Force Microscopy (AFM), which can both sense and apply forces at the nanoscale, with sub-nanonewton sensitivity, represents an enabling technology and a crucial experimental tool in biophysics and mechanobiology. In this work, we report on the application of AFM to the study of biomechanical fingerprints of different components of biological systems, such as the ECM, the whole cell, and cellular components, such as the nucleus, lamellipodia and the glycocalyx. We show that physical observables such as the (spatially resolved) Young’s Modulus (YM) of elasticity of ECMs or cells, and the effective thickness and stiffness of the glycocalyx, can be quantitatively characterized by AFM. Their modification can be correlated to changes in the microenvironment, physio-pathological conditions, or gene regulation.