title: Three-dimensional perfused humanin vitromodel of non-alcoholic fatty liver disease.
abstract: OBJECTIVE
  To develop a humanin vitromodel of non-alcoholic fatty liver disease (NAFLD), utilising primary hepatocytes cultured in a three-dimensional (3D) perfused platform.
  
  
  METHODS
  Fat and lean culture media were developed to directly investigate the effects of fat loading on primary hepatocytes cultured in a 3D perfused culture system. Oil Red O staining was used to measure fat loading in the hepatocytes and the consumption of free fatty acids (FFA) from culture medium was monitored. Hepatic functions, gene expression profiles and adipokine release were compared for cells cultured in fat and lean conditions. To determine if fat loading in the system could be modulated hepatocytes were treated with known anti-steatotic compounds.
  
  
  RESULTS
  Hepatocytes cultured in fat medium were found to accumulate three times more fat than lean cells and fat uptake was continuous over a 14-d culture. Fat loading of hepatocytes did not cause any hepatotoxicity and significantly increased albumin production. Numerous adipokines were expressed by fatty cells and genes associated with NAFLD and liver disease were upregulated including: Insulin-like growth factor-binding protein 1, fatty acid-binding protein 3 and CYP7A1. The metabolic activity of hepatocytes cultured in fatty conditions was found to be impaired and the activities of CYP3A4 and CYP2C9 were significantly reduced, similar to observations made in NAFLD patients. The utility of the model for drug screening was demonstrated by measuring the effects of known anti-steatotic compounds. Hepatocytes, cultured under fatty conditions and treated with metformin, had a reduced cellular fat content compared to untreated controls and consumed less FFA from cell culture medium.
  
  
  CONCLUSIONS
  The 3Din vitroNAFLD model recapitulates many features of clinical NAFLD and is an ideal tool for analysing the efficacy of anti-steatotic compounds.
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Tomasz
authors.initials: T
authors.lastname: Kostrzewski
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Terri
authors.initials: T
authors.lastname: Cornforth
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Sophie A
authors.initials: SA
authors.lastname: Snow
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Larissa
authors.initials: L
authors.lastname: Ouro-Gnao
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Cliff
authors.initials: C
authors.lastname: Rowe
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: Emma M
authors.initials: EM
authors.lastname: Large
authors.affiliation: Tomasz Kostrzewski, Terri Cornforth, Sophie A Snow, Larissa Ouro-Gnao, Cliff Rowe, Emma M Large, David J Hughes, CN Bio Innovations Limited, Welwyn Garden City AL7 3AX, United Kingdom.
authors.forename: David J
authors.initials: DJ
authors.lastname: Hughes
mesh_terms: D017667:Adipocytes / Q000378:metabolism*; D054392:Adipokines / Q000378:metabolism*; D001391:Azo Compounds / Q000008:administration & dosage; D019149:Bioreactors; D018929:Cell Culture Techniques; D002790:Cholesterol 7-alpha-Hydroxylase / Q000378:metabolism; D004396:Coloring Agents / Q000008:administration & dosage; D015925:Cryopreservation; D065729:Cytochrome P-450 CYP2C9 / Q000378:metabolism; D051544:Cytochrome P-450 CYP3A / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005230:Fatty Acids, Nonesterified / Q000378:metabolism*; D022781:Hepatocytes / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D018970:Insulin-Like Growth Factor Binding Protein 1 / Q000378:metabolism; D008954:Models, Biological; D065626:Non-alcoholic Fatty Liver Disease / Q000378:metabolism*; D010477:Perfusion; D061251:Primary Cell Culture; D054457:Tissue Scaffolds; D014280:Triglycerides / Q000378:metabolism
chemical_list: D054392:Adipokines; D001391:Azo Compounds; D004396:Coloring Agents; D005230:Fatty Acids, Nonesterified; C032231:IGFBP1 protein, human; D018970:Insulin-Like Growth Factor Binding Protein 1; D014280:Triglycerides; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D051544:Cytochrome P-450 CYP3A; C553219:CYP7A1 protein, human; D002790:Cholesterol 7-alpha-Hydroxylase; C510163:CYP3A4 protein, human; C011049:oil red O
keywords: Fatty liver; Hepatocytes; Liver disease; Non-alcoholic fatty liver disease; Organ-on-chip; Primary cell culture; Three-dimensional cell culture