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T-cadherin modulates hepatocyte functions in vitro.

ABSTRACT: Primary hepatocytes from several different species rapidly lose viability and phenotypic functions on isolation from their native microenvironment of the liver. Stromal cells derived from both within and outside the liver can induce phenotypic functions in primary hepatocytes in vitro; however, the molecular mediators underlying this "coculture effect" have not been fully elucidated. We have previously developed a functional genomic screen utilizing cocultures of hepatocytes and 3T3 fibroblasts to identify such candidate hepatocyte-function-inducing molecules. In particular, truncated-cadherin (T-cadherin) was identified as a potential molecule of interest in induction of hepatic functions. Here we demonstrate that liver-specific functions of primary rat hepatocytes are induced on cocultivation with Chinese hamster ovary cells engineered to express T-cadherin on their surface as compared with wild-type controls. Additionally, culture of cells on substrata presenting recombinant T-cadherin protein (acellular presentation) enhanced hepatic functions in both pure hepatocyte cultures and in hepatocyte-stromal cocultures lacking endogenous T-cadherin expression. Collectively, these data indicate that both cellular and acellular presentation of T-cadherin can be used to modulate the hepatocyte phenotype in vitro for tissue engineering applications. Our work suggests potential avenues for investigating the role of T-cadherin on hepatocellular function in vivo in settings such as embryogenesis and liver pathology.

SUBMITTER: Khetani SR

PROVIDER: S-EPMC2574031 | biostudies-literature | 2008 Nov

REPOSITORIES: biostudies-literature

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T-cadherin modulates hepatocyte functions in vitro.

Khetani Salman R SR Chen Alice A AA Ranscht Barbara B Bhatia Sangeeta N SN

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20080717 11

Primary hepatocytes from several different species rapidly lose viability and phenotypic functions on isolation from their native microenvironment of the liver. Stromal cells derived from both within and outside the liver can induce phenotypic functions in primary hepatocytes in vitro; however, the molecular mediators underlying this "coculture effect" have not been fully elucidated. We have previously developed a functional genomic screen utilizing cocultures of hepatocytes and 3T3 fibroblasts ...[more]

PMID: 18635739

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Project description:Liver fibrosis occurs as a consequence of chronic injuries from viral infections, metabolic disorders, and alcohol abuse. Fibrotic liver microenvironment (LME) is characterized by excessive deposition and aberrant turnover of extracellular matrix proteins, which leads to increased tissue stiffness. Liver stiffness acts as a vital cue in the regulation of hepatic responses in both healthy and diseased states; however, the effect of varying stiffness on liver cells is not well understood. There is a critical need to engineer in vitro models that mimic the liver stiffness corresponding to various stages of disease progression in order to elucidate the role of individual cellular responses. Here we employed polydimethyl siloxane (PDMS) based substrates with tunable mechanical properties to investigate the effect of substrate stiffness on the behavior of primary rat hepatocytes. To recreate physiologically relevant stiffness, we designed soft substrates (2 kPa) to represent the healthy liver and stiff substrates (55 kPa) to represent the diseased liver. Tissue culture plate surface (TCPS) served as the control substrate. We observed that hepatocytes cultured on soft substrates displayed a more differentiated and functional phenotype for a longer duration as compared to stiff substrates and TCPS. We demonstrated that hepatocytes on soft substrates exhibited higher urea and albumin synthesis. Cytochrome P450 (CYP) activity, another critical marker of hepatocytes, displayed a strong dependence on substrate stiffness, wherein hepatocytes on soft substrates retained 2.7 fold higher CYP activity on day 7 in culture, as compared to TCPS. We further observed that an increase in stiffness induced downregulation of key drug transporter genes (NTCP, UGT1A1, and GSTM-2). In addition, we observed that the epithelial cell phenotype was better maintained on soft substrates as indicated by higher expression of hepatocyte nuclear factor 4?, cytokeratin 18, and connexin 32. These results indicate that the substrate stiffness plays a significant role in modulating hepatocyte behavior. Our PDMS based liver model can be utilized to investigate the signaling pathways mediating the hepatocyte-LME communication to understand the progression of liver diseases.

Dataset Information

T-cadherin modulates hepatocyte functions in vitro.

Publications

T-cadherin modulates hepatocyte functions in vitro.

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