Anisotropic expansion of hepatocyte lumina enforced by apical bulkheads

Abstract

Lumen morphogenesis results from the interplay between molecular pathways and mechanical forces. In several organs, epithelial cells share their apical surfaces to form a tubular lumen. In the liver, however, hepatocytes share the apical surface only between adjacent cells and form narrow lumina that grow anisotropically, generating a 3D network of bile canaliculi (BC). Here, by studying lumenogenesis in differentiating mouse hepatoblasts in vitro, we discovered that adjacent hepatocytes assemble a pattern of specific extensions of the apical membrane traversing the lumen and ensuring its anisotropic expansion. These previously unrecognized structures form a pattern, reminiscent of the bulkheads of boats, also present in the developing and adult liver. Silencing of Rab35 resulted in loss of apical bulkheads and lumen anisotropy, leading to cyst formation. Strikingly, we could reengineer hepatocyte polarity in embryonic liver tissue, converting BC into epithelial tubes. Our results suggest that apical bulkheads are cell-intrinsic anisotropic mechanical elements that determine the elongation of BC during liver tissue morphogenesis.

Hepatocyte apical bulkheads provide a mechanical means to oppose bile pressure

“…During hepatoblast-to-hepatocyte differentiation in the developing liver, adjacent hepatoblasts initially form small spherical lumina between their apical membranes. These apical lumina then expand anisotropically, generating narrow tubules that ultimately connect to complete the formation of the BC network ( Belicova et al, 2021 ). Similar to lumen expansion in other epithelia, BC elongation is driven by secretion of ions (bile acids and other organic solutes) into the nascent lumen ( Boyer, 2013 ; Bryant and Mostov, 2008 ; Dasgupta et al, 2018 ).…”

“…To achieve anisotropic expansion of the apical lumen, hepatocytes need to balance the isotropic hydrostatic pressure in the lumen with anisotropic forces in the subapical actomyosin cortex ( Li et al, 2016 ). We recently reported that BC elongation is enforced by apical bulkheads, F-actin–containing extensions of the apical membrane that traverse the canalicular lumen and connect juxtaposed hepatocytes ( Belicova et al, 2021 ). The structure of apical bulkheads and their role in BC formation suggest that they are mechanical elements that generate anisotropy in the subapical actomyosin cortex, thereby driving BC elongation.…”

“…In this study, we experimentally tested the function of apical bulkheads as mechanical elements that oppose canalicular bile pressure. As a model system, we used in vitro–differentiated hepatocytes that form elongated BC ( Belicova et al, 2021 ). In this culture system, the BC remain closed, preventing the drainage of bile, reflecting the elevated bile pressure that hepatocytes experience during development and in cholestatic liver.…”

Rab8, Rab11, and Rab35 coordinate lumen and cilia formation during Zebrafish Left-Right Organizer development

“…In vitro cell culture assays have been used to identify regulators of lumen establishment or ciliogenesis and have identified 3 potential Rab GTPases that are involved in both: Rab11, Rab8, and Rab35 [18][19][20][21][22][23][24]. Here we investigated the role of these three Rab GTPases in KV development.…”

Rab35 controls formation of luminal projections required for bile canalicular morphogenesis

“…Several pathways have been proposed to play key roles in BC morphogenesis such as mitosis, cytokinesis and endosomal trafficking ( 4 , 5 , 6 ), nevertheless the mechanistic studies are lacking. Belicova and colleagues set out to investigate the basic mechanisms for BC formation ( 7 ).…”