Remodelling of the fibre-aggregate structure of collagen gels by cancer-associated fibroblasts: a time-resolved grey-tone image analysis based on stochastic modelling

Abstract

Solid tumors consist of tumor cells associated with stromal and immune cells, secreted factors and extracellular matrix (ECM), that together constitute the tumor microenvironment. Among stromal cells, cancer-associated fibroblasts (CAFs) are of particular interest. CAFs modulate the architecture of the ECM by exerting forces and contracting collagen fibres, creating paths that facilitate cancer cell migration. The characterization of the collagen fibre network and its space and time-dependent microstructural modifications is key to investigating the interactions between cells and the ECM. Developing image analysis tools for that purpose is still a challenge. The structural complexity of the collagen network calls for specific statistical descriptors. Moreover, the low signalto-noise ratio of imaging techniques available for time-resolved studies rules out standard image analysis methods. In this work, we propose an innovative data analysis approach to investigate the remodelling of fibrillar collagen in a three-dimensional spheroid model of cellular invasion, which is based on the stochastic modelling of the gel structure and on grey-tone image analysis. The method was used to study the reshaping of a collagen matrix by migrating breast cancer-derived CAFs. The structure of the gels at the scale of a few microns consists in regions with high fibre density separated by depleted regions, which can be thought of as fibre aggregates and pores. To characterize this structure, we developed a two-scale stochastic model with a clipped Gaussian field model to describe the aggregates-and-pores large-scale structure, and a homogeneous Boolean model to describe the small-scale fibre network within the aggregates. The model parameters are identified by fitting the grey-tone histograms and correlation functions of confocal microscopy images. The specificity of the method is that it applies to the unprocessed grey-tone images, and it can therefore be used with low magnification, noisy time-lapse reflectance images. When applied to the spheroid time-resolved images, the method revealed two different matrix densification mechanisms for the gel in direct contact or far from the cells.