title: New physiologically-relevant liver tissue model based on hierarchically cocultured primary rat hepatocytes with liver endothelial cells.
abstract: To develop an in vitro liver tissue equivalent, hepatocytes should be cocultured with liver non-parenchymal cells to mimic the in vivo physiological microenvironments. In this work, we describe a physiologically-relevant liver tissue model by hierarchically organizing layers of primary rat hepatocytes and human liver sinusoidal endothelial cells (TMNK-1) on an oxygen-permeable polydimethylsiloxane (PDMS) membrane, which facilitates direct oxygenation by diffusion through the membrane. This in vivo-mimicking hierarchical coculture was obtained by simply proceeding the overlay of TMNK-1 cells on the hepatocyte layer re-formed on the collagen immobilized PDMS membranes. The comparison of hepatic functionalities was achieved between coculture and sandwich culture with Matrigel, in the presence and absence of direct oxygenation. A complete double-layered structure of functional liver cells with vertical contact between hepatocytes and TMNK-1 was successfully constructed in the coculture with direct oxygen supply and was well-maintained for 14 days. The hepatocytes in this hierarchical culture exhibited improved survival, functional bile canaliculi formation, cellular level polarization and maintenance of metabolic activities including Cyp1A1/2 activity and albumin production. By contrast, the two cell populations formed discontinuous monolayers on the same surfaces in the non-oxygen-permeable cultures. These results demonstrate that (i) the direct oxygenation through the PDMS membranes enables very simple formation of a hierarchical structure consisting of a hepatocyte layer and a layer of TMNK-1 and (ii) we may include other non-parenchymal cells in this format easily, which can be widely applicable to other epithelial organs.
authors: Fe505, Institute of Industrial Science (IIS), University of Tokyo, Tokyo, 153-8505, Japan. w-xiao@iis.u-tokyo.ac.jp.
authors: Wenjin
authors: W
authors: Xiao
authors: Fe505, Institute of Industrial Science (IIS), University of Tokyo, Tokyo, 153-8505, Japan. w-xiao@iis.u-tokyo.ac.jp and LIMMS CNRS/IIS, University of Tokyo, Japan.
authors: Guillaume
authors: G
authors: Perry
authors: Fe505, Institute of Industrial Science (IIS), University of Tokyo, Tokyo, 153-8505, Japan. w-xiao@iis.u-tokyo.ac.jp.
authors: Kikuo
authors: K
authors: Komori
authors: Fe505, Institute of Industrial Science (IIS), University of Tokyo, Tokyo, 153-8505, Japan. w-xiao@iis.u-tokyo.ac.jp.
authors: Yasuyuki
authors: Y
authors: Sakai
mesh_terms: D000418:Albumins / Q000737:chemistry; D000818:Animals; D001646:Bile / Q000737:chemistry; D018929:Cell Culture Techniques / Q000379:methods; D002462:Cell Membrane / Q000378:metabolism; D018920:Coculture Techniques; D004058:Diffusion; D004129:Dimethylpolysiloxanes / Q000737:chemistry; D042783:Endothelial Cells / Q000166:cytology*; D005434:Flow Cytometry; D022781:Hepatocytes / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008099:Liver / Q000473:pathology*; D008297:Male; D008567:Membranes, Artificial; D010100:Oxygen / Q000737:chemistry; D051381:Rats; D017208:Rats, Wistar; D023822:Tissue Engineering
chemical_list: D000418:Albumins; D004129:Dimethylpolysiloxanes; D008567:Membranes, Artificial; C013830:baysilon; D010100:Oxygen