Structural Polarity and Functional Bile Canaliculi in Rat Hepatocyte Spheroids

Abu-Absi, Susan Fugett; Friend, Julie R.; Hansen, Linda K.; Hu, Wei Shou

doi:10.1006/excr.2001.5467

Cited by 218 publications

(174 citation statements)

References 49 publications

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“…At least part of the interaction between hepatocytes and LSECs has been shown to be mediated by growth factors. 29,47 Similarly, hepatocyte spheroids were shown to express E-cadherin, 34 form extensive bile canaliculi, 48 and show sinusoidal surface markers at the interface between cells in the spheroid. 48 In conclusion, cocultures of LSECs and hepatocytes express high levels of LDL-R and therefore could be used for the study of LDL uptake and clearance by primary hepatocytes.…”

Section: Discussionmentioning

confidence: 99%

Liver endothelial cells promote LDL-R expression and the uptake of HCV-like particles in primary rat and human hepatocytes

et al. 2006

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Low-density lipoprotein (LDL) is an important carrier of plasma cholesterol and triglycerides whose concentration is regulated by the liver parenchymal cells. Abnormal LDL regulation is thought to cause atherosclerosis, while viral binding to LDL has been suggested to facilitate hepatitis C infection. Primary hepatocytes quickly lose the ability to clear LDL during in vitro culture. Here we show that the coculture of hepatocytes with liver sinusoidal endothelial cells (LSEC) significantly increases the ability of hepatocytes to uptake LDL in vitro. LDL uptake does not increase when hepatocytes are cocultured with other cell types such as fibroblasts or umbilical vein endothelial cells. We find that LSECs induce the hepatic expression of the LDL receptor and the epidermal growth factor receptor. In addition, while hepatocytes in single culture did not take up hepatitis C virus (HCV)-like particles, the hepatocytes cocultured with LSECs showed a high level of HCV-like particle uptake. We suggest that coculture with LSECs induces the emergence of a sinusoidal surface in primary hepatocytes conducive to the uptake of HCV-like particles. In conclusion, our findings describe a novel model of polarized hepatocytes in vitro that can be used for the study of LDL metabolism and hepatitis C infection. L ow-density lipoprotein (LDL) is a plasma-carried particle whose lipid component includes cholesterol and triglycerides. 1 LDL originates from very low-density lipoprotein (vLDL) synthesized by the liver with apoprotein B-100. The vLDL is converted to LDL by an endothelial lipase, which releases free fatty acids, increasing the density of the particle to form LDL. Excess LDL is then taken up by hepatocytes through LDL receptor (LDL-R)-mediated endocytosis. 1 Improper hepatic clearance of LDL results in elevated plasma levels of LDL which is a serious risk factor for the development of atherosclerosis. 2 One of the first steps in atherosclerosis is the passage of LDL into the vascular wall. Once trapped in the vessel wall, LDL can undergo oxidation. 3 Oxidized LDL is no longer a ligand for the LDL receptor. 4 Accumulation of oxidized LDL and LDL aggregates in the vessel wall stimulates an inflammatory response causing endothelial injury, macrophage recruitment, foam cell formation, and smooth muscle cell proliferation. 3 Current therapeutic intervention includes administration of statins, which block cholesterol production and increase expression of the LDL-R by the liver parenchymal cells, 2 removing LDL from circulation. 5 In addition, liver sinusoidal endothelial cells (LSECs) remove oxidized LDL from the circulation via the scavenger cell receptor. 5 LDL-and LDL-R-mediated pathways have also been shown to play a role in hepatitis C virus (HCV) infection. 6 The HCV surface receptors, glycoproteins E1 and E2, have been shown to associate with LDL and vLDL. 7 The virus particle itself was shown to associate with the

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Section: Discussionmentioning

confidence: 99%

Liver endothelial cells promote LDL-R expression and the uptake of HCV-like particles in primary rat and human hepatocytes

et al. 2006

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“…Some of these techniques result in the hepatocytes forming multicellular spheroids. The structure of the spheroids appears to mimic, in some respects, that of liver tissue in vivo -for example, channels resembling bile canaliculi are seen (Abu-Absi et al, 2002). Spheroid culture also results in prolonged expression of liverspecific functions (commonly measured by albumin production (Riccalton-Banks, 2002)) and hepatocyte viability (Richert et al, 2002).…”

Section: Introductionmentioning

confidence: 98%

A Mathematical Model of Liver Cell Aggregation In Vitro

et al. 2008

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"Authors may self-archive the author's accepted manuscript of their articles on their own websites. Authors may also deposit this version of the article in any repository, provided it is only made publicly available 12 months after official publication or later. He/ she may not use the publisher's version (the final article), which is posted on SpringerLink and other Springer websites, for the purpose of self-archiving or deposit. Furthermore, the author may only post his/her version provided acknowledgement is given to the original source of publication and a link is inserted to the published article on Springer's website. The link must be accompanied by the following text: "The final publication is available at link.springer.com"". th March, 2014A mathematical model of liver cell aggregation in vitroThe behaviour of mammalian cells within three-dimensional structures is an area of intense biological research and underpins the efforts of tissue engineers to regenerate human tissues for clinical applications. In the particular case of hepatocytes (liver cells), the formation of spheroidal multicellular aggregates has been shown to improve cell viability and functionality compared to traditional monolayer culture techniques.We propose a simple mathematical model for the early stages of this aggregation process, when cell clusters form on the surface of the extracellular matrix (ECM) layer on which they are seeded. We focus on interactions between the cells and the viscoelastic ECM substrate. Governing equations for the cells, culture medium and ECM are derived using the principles of mass and momentum balance. The model is then reduced to a system of four partial differential equations, which are investigated analytically and numerically. The model predicts that, provided cells are seeded at a suitable density, aggregates with clearly defined boundaries and a spatially uniform cell density on the interior will form. While the mechanical properties of the ECM do not appear to have a significant effect, strong cell-ECM interactions can inhibit, or possibly prevent, the formation of aggregates. The paper concludes with a discussion of our key findings and suggestions for future work.

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“…Cell culture media was purchased from Invitrogen (Carlsbad, CA), [1,[2][3] H]-cortisone from American Radiolabeled Chemicals (St.Louis, MO), and all other chemicals were from Fluka AG (Buchs, Switzerland).…”

Section: Methodsmentioning

confidence: 99%

“…Although primary hepatocytes cultured as a monolayer are commonly used, their application is restricted to the assessment of acute toxicity due to limited viability, altered polarity, rapid loss of liver-specific gene expression and rapidly decreasing functionality [2,3]. Hepatocytes cultured as two-dimensional (2D) monolayers often fail to predict chronic toxicity, and only 50% of acute hepatotoxic compounds are detected [4].…”

Section: Introductionmentioning

confidence: 99%

Characterization of a Rat Multi-Cell Type 3D-Liver Microtissue System

Odermatt¹

2013

J Tissue Sci Eng

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Background: Primary hepatocytes rapidly lose their polarized morphology and the expression of important liverspecific metabolic enzymes, receptors and transport proteins under normal two-dimensional (2D) culture conditions. Thus, their use as a reliable predictive in vitro model for drug-induced liver injury is limited. Three-dimensional (3D) liver micro tissue culture systems have become an increasingly attractive alternative for the evaluation of druginduced liver toxicity. However, several liver-specific pathways remain to be characterized in such models. Methods and principal findings:In the present work, we compared the expression of several genes with a role in the anti-oxidant cell defense and glucocorticoid pathways in rat H4IIE hepatoma cells, primary rat hepatocytes, 2D-hepatocyte sandwich culture, and a multi-cell type micro tissue model comprising primary rat hepatocytes in coculture with liver-derived nonparenchymal cells and macrophage (Kupffer cells). Gene expression was studied for up to 25 days in culture. High expression levels of the Nrf2-dependent genes NQO1, ABCC3 and GST2A were detected in H4IIE cells and in 3D-liver microtissues, in contrast to 2D-hepatocytes where a rapid decline of these genes was observed. The glucocorticoid-dependent genes ORM1, G6PC, PCK1 and HSD11B1 were highly expressed up to 25 days of cultivation in 3D-liver microtissues, but they showed very low or background expression in H4IIE and 2D-hepatocyte models. Conclusions:The tested 3D-multi-cell type liver micro tissue represents a stable and functionally active model system, with sustained expression for more than three weeks of cultivation of important metabolic proteins regulated by the glucocorticoid and anti-oxidant cell defense pathway.

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Structural Polarity and Functional Bile Canaliculi in Rat Hepatocyte Spheroids

Cited by 218 publications

References 49 publications

Liver endothelial cells promote LDL-R expression and the uptake of HCV-like particles in primary rat and human hepatocytes

Liver endothelial cells promote LDL-R expression and the uptake of HCV-like particles in primary rat and human hepatocytes

A Mathematical Model of Liver Cell Aggregation In Vitro

Characterization of a Rat Multi-Cell Type 3D-Liver Microtissue System

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