P-5 three-dimensional single-cell atlas of liver tissue architecture

Introduction and Objectives: The liver is an organ that performs a wide variety of functions that are highly dependent on its complex 3D structure. Geometrical models (digital representations of tissues) represent a versatile technique to characterize 3D tissues as well as to get quantitative insights into the link between their structure and function. Until now, these models have only focused on some tissue (sinusoids and bile canaliculi) and cellular components (hepatocytes), leaving out important cellular populations such as stellate cells and Kupffer cells. One of the major bottlenecks for a complete tissue reconstruction is the limitation on the number of markers that can be imaged by fluorescence microscopy (up to 4-5). This study aimed to generate a “3D single-cell atlas of liver tissue architecture”, i.e., a full 3D geometrical model which includes all tissue and cellular components simultaneously. Materials and Methods: We overcome the technical constraints by using deep tissue immunostaining, multiphoton microscopy, deep learning techniques, and 3D image processing. As a proof of concept, we used the 3D atlas to describe the morphological changes that occur in the mouse liver during post-natal early development and adulthood. Results: We described how liver tissue architecture progressed from post-natal day one to adulthood by a novel set of morphometric cellular and tissue parameters. Our analysis revealed unknown details about the spatial organization of different liver cell types. Unexpected spatiotemporal patterns of non-parenchymal cells and hepatocytes with differing in size, number of nuclei, and DNA content were uncovered. We also provided information regarding the remodeling of the bile canaliculi and sinusoidal networks. Conclusions: These findings revealed novel characteristics of liver heterogeneity and have important implications for both the structural organization of liver tissue and its functional features. 3D single-cell atlas will provide a powerful tool to understand liver tissue architecture under both physiological and pathological conditions.