Fat-loaded HepG2 spheroids exhibit enhanced protection from Pro-oxidant and cytokine induced damage.
The mechanisms by which steatosis renders hepatocytes susceptible to damage in non-alcoholic steatohepatitis (NASH) are unclear although fat accumulation is believed to increase hepatocyte susceptibility to inflammatory cytokines and oxidative stress. We therefore investigated the susceptibility of steatotic, hepatocyte-derived cells to TNFalpha and the pro-oxidant, t-butylhydroperoxide (TBH). HepG2 spheroids rendered steatotic by fat-loading with 0.15 mM oleic or palmitic acid for 48 h and treated with TNFalpha or TBH for 18 h exhibited surprisingly lower levels of cytotoxicity, and increased anti-oxidant activity (superoxide dismutase (SOD)) compared with non fat-loaded controls. The protective effect of steatosis was significantly reversed by the inhibition of AMP-activated kinase (AMPK) since spheroids transfected with a kinase-dead AMPKalpha2 subunit, exhibited a significant increase in TBH-induced cytotoxicity when fat-loaded. In conclusion, our findings suggest that fat-loaded hepatocyte-derived cells are surprisingly less susceptible to cytokine and pro-oxidant induced damage via an adaptive mechanism dependent, in part, on AMPK activity.
Damelin
Leonard H
LH
UCL Institute of Hepatology, Hampstead Campus, Rowland Hill Street, London NW3 2PF, United Kingdom. l.damelin@medsch.ucl.ac.uk
Coward
Sam
S
Kirwan
Michael
M
Collins
Peter
P
Selden
Clare
C
Hodgson
Humphrey J F
HJ
D055372:AMP-Activated Protein Kinases; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006801:Humans; D050356:Lipid Metabolism / Q000187:drug effects; D046529:Microscopy, Electron, Transmission; D009097:Multienzyme Complexes / Q000378:metabolism; D019301:Oleic Acid / Q000378:metabolism* / Q000494:pharmacology; D010084:Oxidation-Reduction / Q000187:drug effects; D018384:Oxidative Stress; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D018874:Spheroids, Cellular / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D013482:Superoxide Dismutase / Q000378:metabolism; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D020122:tert-Butylhydroperoxide / Q000494:pharmacology*
D009097:Multienzyme Complexes; D017392:Thiobarbituric Acid Reactive Substances; D014409:Tumor Necrosis Factor-alpha; D019301:Oleic Acid; D020122:tert-Butylhydroperoxide; D013482:Superoxide Dismutase; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases