An effective protocol for Pharmacological Defatting of Primary Human Hepatocytes which is non-toxic to Cholangiocytes or Intrahepatic endothelial cells

Research output: Contribution to journalArticle


  • Amanda Pinter Carvalheiro da Silva Boteon
  • Darius Mirza
  • H Mergentel

Colleges, School and Institutes

External organisations

  • National Institute for Health Research Liver Biomedical Research Unit, University Hospitals Birmingham NHS Foundation Trust and the University of Birmingham, Birmingham, UK; Centre for Liver Research, Institute of Immunology and Immunotherapy, University of Birmingham, Birmingham, UK.
  • Liver Unit, University Hospitals Birmingham NHS Foundation Trust, Birmingham, UK.
  • National Institute for Health Research (NIHR) Birmingham Biomedical Research Centre, University Hospitals Birmingham NHS Foundation Trust and University of Birmingham, Birmingham, UK


Introduction: Pharmacological defatting of rat hepatocytes and hepatoma cell lines suggests that the same method could be used to ameliorate macrovesicular steatosis in moderate to severely fatty livers. However there is no data assessing the effects of those drugs on primary human liver cells. We aimed to determine the effectiveness of a pharmacological cocktail in reducing the in vitro lipid content of primary human hepatocytes (PHH). In addition we sought to determine the cytotoxicity of the cocktail towards non-parenchymal liver cells. Methods: Steatosis was induced in PHH by supplementation with a combination of saturated and unsaturated free fatty acids. This was followed by addition of a defatting drug cocktail for up to 48 hours. The same experimental method was used with human intra-hepatic endothelial cells (HIEC) and human cholangiocytes. MTT assay was used to assess cell viability, triglyceride quantification and oil red O staining were used to determine intracellular lipids content whilst ketone bodies were measured in the supernatants following experimentation. Results: Incubation of fat loaded PHH with the drugs over 48 hours reduced the intracellular lipid area by 54%, from 12.85% to 5.99% (p=0.002) (percentage of total oil red O area), and intracellular triglyceride by 35%, from 28.24 to 18.30 nmol/million of cells (p<0.001). Total supernatant ketone bodies increased 1.4-fold over 48 hours in the defatted PHH compared with vehicle controls (p=0.002). Moreover incubation with the drugs for 48 hours increased the viability of PHH by 11%, cholangiocytes by 25% whilst having no cytotoxic effects on HIEC. Conclusion: These data demonstrate that pharmacological intervention can significantly decrease intracellular lipid content of PHH, increase fatty acids β-oxidation whilst being non-toxic to PHH, HIEC or cholangiocytes.


Original languageEnglish
Article numbere0201419
JournalPLoS ONE
Issue number7
Publication statusPublished - 25 Jul 2018


  • defatting, Primary Human Hepatocytes, liver transplantation, steatosis

ASJC Scopus subject areas