Abstract
Human stem cell-derived hepatocyte-like cells (HLCs) offer an attractive platform to study liver biology. Despite their numerous advantages, HLCs lack critical in vivo characteristics, including cell polarity. Here, we report a stem cell differentiation protocol that uses transwell filters to generate columnar polarized HLCs with clearly defined basolateral and apical membranes separated by tight junctions. We show that polarized HLCs secrete cargo directionally: Albumin, urea, and lipoproteins are secreted basolaterally, whereas bile acids are secreted apically. Further, we show that enterically transmitted hepatitis E virus (HEV) progeny particles are secreted basolaterally as quasi-enveloped particles and apically as naked virions, recapitulating essential steps of the natural infectious cycle in vivo. We also provide proof-of-concept that polarized HLCs can be used for pharmacokinetic and drug-drug interaction studies. This novel system provides a powerful tool to study hepatocyte biology, disease mechanisms, genetic variation, and drug metabolism in a more physiologically relevant setting.
Highlights
Human stem cell-derived hepatocyte-like cells (HLCs) offer an attractive platform to study liver biology
Maintenance of cell polarity is essential for retaining primary human hepatocytes (PHH) functions, yet, HLCs are conventionally differentiated in culture dishes under two-dimensional (2D) culture conditions, a process that is inefficient, variable, and yields lesspolarized cells
When HLCs are differentiated in spheroids[8,9] or cultured in micropatterned cocultures[10] they better recapitulate hepatocyte functions
Summary
Human stem cell-derived hepatocyte-like cells (HLCs) offer an attractive platform to study liver biology. Hepatocytes form a crucial cell layer engaged in two counter-current flow systems, which, on the one hand, involve uptake, processing, and secretion of sinusoidal blood components, and on the other hand, synthesis and secretion of bile[1] To mediate these functions, hepatocytes have a unique polarization with multiple basolateral membranes facing the sinusoids, and multiple apical membranes forming bile canaliculi (Fig. 1a, right panel). Hepatocytes have a unique polarization with multiple basolateral membranes facing the sinusoids, and multiple apical membranes forming bile canaliculi (Fig. 1a, right panel) Within this peculiar structure, cell signaling, membrane trafficking, protein secretion, and bile transport are highly organized[2]. Hepatocyte-based research would benefit from more reliable, physiologically relevant, and more experimentally tractable hepatocellular polarity systems
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