Quantification of topological features in cell meshes to explore E-cadherin dysfunction

Abstract

In cancer, defective E-cadherin leads to cell detachment, migration and metastization. Further, alterations mediated by E-cadherin dysfunction affect cell topology and tissue organization. Herein, we propose a novel quantitative approach, based on microscopy images, to analyse abnormal cellular distribution patterns. We generated undirected graphs composed by sets of triangles which accurately reproduce cell positioning and structural organization within each image. Network analysis was developed by exploring triangle geometric features, namely area, edges length and formed angles, as well as their variance, when compared with the respective equilateral triangles. We generated synthetic networks, mimicking the diversity of cell-cell interaction patterns, and evaluated the applicability of the selected metrics to study topological features. Cells expressing wild-type E-cadherin and cancer-related mutants were used to validate our strategy. Specifically, A634V, R749W and P799R cancer-causing mutants present more disorganized spatial distribution when compared with wild-type cells. Moreover, P799R exhibited higher length and angle distortions and abnormal cytoskeletal organization, suggesting the formation of very dynamic and plastic cellular interactions. Hence, topological analysis of cell network diagrams is an effective tool to quantify changes in cell-cell interactions and, importantly, it can be applied to a myriad of processes, namely tissue morphogenesis and cancer.

Predicting the Functional Impact of CDH1 Missense Mutations in Hereditary Diffuse Gastric Cancer

“…Importantly, functional impairment of E-cadherin and eventual loss of the molecule is typically associated with decreased cell-cell interaction and tissue remodelling [ 3 , 5 , 6 ]. Recently, to characterize defects in the epithelial structure and morphology that can arise from E-cadherin mutants, we developed a platform that identifies and quantifies cellular distribution patterns using in situ microscopy images [ 46 ]. More specifically, we used DAPI-stained nuclei to create artificial cellular networks, from which we could extract quantitative data regarding cell distribution and organization ( Figure 3 A).…”

“…More specifically, we used DAPI-stained nuclei to create artificial cellular networks, from which we could extract quantitative data regarding cell distribution and organization ( Figure 3 A). The software creates digital meshes composed of triangles centred in triplets of neighbouring nuclei, and explores their topological features, such as area, edges length, and angles [ 46 ]. Pathogenic mutations with impact in cellular organization, present triangles with higher areas and edges when compared with the wild-type cell counterparts.…”

Blue intensity matters for cell cycle profiling in fluorescence DAPI-stained images

“…38 These are, undoubtedly, relevant aspects to be consider when using this framework for histological analysis, as the intrinsic characteristics of this imaging framework derive from area and total intensity of DAPI-stained nuclei. Nevertheless, our segmentation algorithm has proven to be very efficient for nuclei or cell segmentations, 13,25 and in fact its semiautomatism enables the operator to critically evaluate and even define nuclei segmentation in histological samples. To perform a comprehensive analysis of more complex samples resorting to this imaging framework, image acquisition procedures should follow the recommendations stated herein.…”

“…13 Applying the above-mentioned criteria, this imaging tool renders the possibility to complement the cell cycle staging with other fluorescence-based multiparametric analyses. This imaging framework can be adapted to different samples as long as the DNA staining protocols enable a quantitative assessment of the DNA based on fluorescence.…”

“…13 The application of these intensity conditioning procedures are crucial for the method proficiency, as both nuclei segmentation and cell cycle phasing depend on two key discriminative features of the stained nucleus, 'area' and 'total intensity'.…”