Microvascular engineering for the development of a non-embedded liver sinusoid with a lumen: when endothelial cells do not lose their edge

Abstract

Vascular engineering seeks to generate reliable models for studying cellular interactions in the context of preserving homeostasis and disease development. However, current systems mainly focus on recapitulating large blood vessels embedded in thick extracellular matrix gels or do not address discontinuous capillaries, crucial microvascular structures. By adapting a Marangoni flow-driven protein densification process on GelTrex domes, we promoted the self-organization of endothelial cells into mature lumenized vessels by anisotropic directional organization following an edge micropattern. Our findings revealed a sequential morphogenetic process leading to microvessels with both murine and human immortalized liver sinusoidal endothelial cells (LSECs). Differential behaviors were found on the whole sample and cell migration, proliferation and polarization were guided by this pattern to form a long multicellular cord. Cells forming this structure deformed large regions of the dome, generating a wave-like fold all around it, hinged in a laminin depletion zone. This wrapped the cell cords with proteins from the gel which marked the beginning of a lumenogenesis process, regulated by a change in the apico-basal polarization of the cord cells, followed by maturation of tight junctions, remodeling of the extracellular matrix, and formation of a lumen; all steps reported inin vivovessel development. Furthermore, we demonstrate that the geometry of the vessels can be controlled by initial topography of the gel. We believe our simple fabrication method, guiding an autonomous self-organization of vessels without the need for supporting cells or complex prefabricated scaffolds, is suitable for future integration into microphysiological systems of discontinuous, fenestrated capillaries.