title: Metabolic Flux Distribution during Defatting of Steatotic Human Hepatoma (HepG2) Cells.
abstract: Methods that rapidly decrease fat in steatotic hepatocytes may be helpful to recover severely fatty livers for transplantation. Defatting kinetics are highly dependent upon the extracellular medium composition; however, the pathways involved are poorly understood. Steatosis was induced in human hepatoma cells (HepG2) by exposure to high levels of free fatty acids, followed by defatting using plain medium containing no fatty acids, or medium supplemented with a cocktail of defatting agents previously described before. We measured the levels of 28 extracellular metabolites and intracellular triglyceride, and fed the data into a steady-state mass balance model to estimate strictly intracellular fluxes. We found that during defatting, triglyceride content decreased, while beta-oxidation, the tricarboxylic acid cycle, and the urea cycle increased. These fluxes were augmented by defatting agents, and even more so by hyperoxic conditions. In all defatting conditions, the rate of extracellular glucose uptake/release was very small compared to the internal supply from glycogenolysis, and glycolysis remained highly active. Thus, in steatotic HepG2 cells, glycolysis and fatty acid oxidation may co-exist. Together, these pathways generate reducing equivalents that are supplied to mitochondrial oxidative phosphorylation.
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. gabriel.yarmush@gmail.com.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Yarmush
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. lucasvcsantos2@gmail.com.
authors.forename: Lucas
authors.initials: L
authors.lastname: Santos
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. josh.yarmush@gmail.com.
authors.forename: Joshua
authors.initials: J
authors.lastname: Yarmush
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. srivaths1992@gmail.com.
authors.forename: Srivathsan
authors.initials: S
authors.lastname: Koundinyan
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. mubasher.saleem1@gmail.com.
authors.forename: Mubasher
authors.initials: M
authors.lastname: Saleem
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. nirnativ1@hotmail.com.
authors.forename: Nir I
authors.initials: NI
authors.lastname: Nativ
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. schloss@soemail.rutgers.edu.
authors.forename: Rene S
authors.initials: RS
authors.lastname: Schloss
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. yarmush@rci.rutgers.edu.|Center for Engineering in Medicine/Surgical Services, Massachusetts General Hospital and the Shriners Hospitals for Children, Boston, MA 02114, USA. yarmush@rci.rutgers.edu.
authors.forename: Martin L
authors.initials: ML
authors.lastname: Yarmush
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. timjm@rci.rutgers.edu.
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Maguire
authors.affiliation: Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854, USA. fberthia@rci.rutgers.edu.
authors.forename: Francois
authors.initials: F
authors.lastname: Berthiaume
keywords: beta-oxidation; defatting; fatty liver; hepatocytes; liver transplantation; mass balances; steatosis