Epithelial organisation revealed by a network of cellular contacts

Abstract

The emergence of differences in the arrangement of cells is the first step towards the establishment of many organs. Understanding this process is limited by the lack of systematic characterization of epithelial organisation. Here we apply network theory at the scale of individual cells to uncover patterns in cell-to-cell contacts that govern epithelial organisation. We provide an objective characterisation of epithelia using network representation, where cells are nodes and cell contacts are links. The features of individual cells, together with attributes of the cellular network, produce a defining signature that distinguishes epithelia from different organs, species, developmental stages and genetic conditions. The approach permits characterization, quantification and classification of normal and perturbed epithelia, and establishes a framework for understanding molecular mechanisms that underpin the architecture of complex tissues.

Figure 1. GNEO approach and epithelial comparisons performed

(a) Geometric and Network representation of Epithelial Organisation (GNEO) approach to characterise epithelial organisation. (1) Images from the confocal microscope are processed to get a light background with dark cell contours. (2) The processed image is the source for defining individual cells, as well as for determining the number of neighbours for every cell. (3) This information is used to produce an epithelial network where each cell is represented as a node and two nodes are connected if the two cells are neighbours in the epithelium. (4) A region of interest (ROI; shown in a green box) is selected for further analysis and the cells that border the ROI are excluded. (5) The average and standard deviation of area and three network features over all cells in an epithelium are calculated. (6) This information is represented as a feature vector, which is an 8-dimensional vector that characterises each epithelium. Each of the four features considered in this work is abbreviated by the symbols shown in this figure. (b) Schematic representation of the comparisons of epithelia from different sources performed in this study. Images of representative epithelial samples (2-pixel wide cell contour) from Drosophila (different tones of green labels, fly) and chicken (different tones of brown labels) are shown. The reference prepupal wing pouch epithelium is shown within a gray box. The text in gray denotes the relationship between epithelia from the different sources that were compared. space: spatially separated epithelia from the same organism; time: temporally separated epithelia from different stages of development; type: different type of epithelia (e.g., squamous and columnar); species: epithelia from different organisms (vertebrate (chick) and invertebrate (Drosophila)) and mutation: mutant epithelia.

Figure 2. Discriminant Analysis of the different epithelia

(a) Colour coded matrix representation of the weights of the features contributing to the observed projection in the Discriminant Analysis of the different comparisons. A higher value (darker color) represents a relatively higher contribution of the feature to the separation. (b-g) Discriminant Analysis graphs of the comparisons of epithelia from different sources. (b) dWP-cNT. (c) cNT-cEE. (d) dWP-cNT-cEE. (e) dWP-dNP. (f) dWP-dWL. (g) dWP-dNP-cNT-cEE.

Figure 3. PCA graphs of the wildtype and mutant epithelia

(a) dWP-dMWP. (b) dWP-dNP-dMWP. (c) dWP-dMWP-cEE. (d) dWP-dMWP-cEE-cNT. dWP: Wing prepupa, cNT: chicken Neural Tube, cEE: chicken Embryonic Ectoderm, dMWP: mutant Wing prepupa.