Hypoxia inducible factor-1 alpha induction by tumour necrosis factor-alpha, but not by toll-like receptor agonists, modulates cellular respiration in cultured human hepatocytes.
OBJECTIVE
Genes encoding for some of the mitochondrial proteins are under the control of the transcriptional factor hypoxia inducible factor-1 alpha (HIF-1 alpha), which can accumulate under normoxic conditions in inflammatory states. The aim of this study was to evaluate the effects of cobalt chloride (CoCl(2), a hypoxia mimicking agent), tumour necrosis factor-alpha (TNF-alpha) and toll-like receptor (TLR) -2, -3 and -4 agonists on HIF-1 alpha accumulation, and further on HIF-1 alpha-mediated modulation of mitochondrial respiration in cultured human hepatocytes.


METHODS
The human hepatoma cell line HepG2 was used in this study. Cells were treated with CoCl(2), TNF-alpha and TLR-2, -3 and -4 agonists. HIF-1 alpha was determined by Western blotting and mitochondrial respiration in stimulated cells by high-resolution respirometry.


RESULTS
CoCl(2), TNF-alpha and TLR agonists induced the expression of HIF-1 alpha in a time-dependent fashion. TNF-alpha and CoCl(2), but not TLR agonists, induced a reduction in complex I-, II- and IV-dependent mitochondrial oxygen consumption. TNF-alpha-associated reduction of cellular oxygen consumption was abolished through inhibition of HIF-1 alpha activity by chetomin (CTM). Pretreatment with cyclosporine A prevented CoCl(2)-induced reduction of complex I- and II-dependent mitochondrial oxygen consumption and TNF-alpha-induced reduction of complex-I-dependent respiration, implicating the involvement of the mitochondrial permeability transition pore openings. TNF-alpha and TLR-2, -3 and -4 agonists induced the expression of vascular endothelial growth factor, which was partially abolished by the blockage of HIF-1 alpha with CTM.


CONCLUSIONS
The data suggest that HIF-1 alpha modulates mitochondrial respiration during CoCl(2) and TNF-alpha stimulation, whereas it has no effect when induced with TLR-2, -3 and -4 agonists.
Regueira
Tomas
T
Department of Intensive Care Medicine, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland.
Lepper
Philipp M
PM
Brandt
Sebastian
S
Ochs
Matthias
M
Vuda
Madhusudanarao
M
Takala
Jukka
J
Jakob
Stephan M
SM
Djafarzadeh
Siamak
S
D053148:Caspase 3 / Q000378:metabolism; D019069:Cell Respiration / Q000187:drug effects*; D003035:Cobalt / Q000494:pharmacology; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D010101:Oxygen Consumption / Q000187:drug effects; D051193:Toll-Like Receptors / Q000819:agonists*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D042461:Vascular Endothelial Growth Factor A / Q000032:analysis
D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D042461:Vascular Endothelial Growth Factor A; D003035:Cobalt; D053148:Caspase 3; C018021:cobaltous chloride