Up-regulation of hypoxia inducible factor 1alpha by isoflurane in Hep3B cells.
BACKGROUND
The volatile anesthetic isoflurane induces hypoxia inducible factor (HIF)-1-responsive genes heme oxygenase 1, inducible nitric oxide synthase, and vascular endothelial growth factor (VEGF) expression. Little is known about the extent to which induction of HIF-1alpha is affected by isoflurane.


METHODS
Hep3B cells were exposed to isoflurane at various concentrations (0.5-4%) or for different time periods (2-8 h) at 37 degrees C. HIF-1alpha gene expression and transcriptional activity, heme oxygenase 1, inducible nitric oxide synthase, and VEGF gene expression were quantified.


RESULTS
Isoflurane induced a time- and concentration-dependent increase in HIF-1alpha protein but not for HIF-1alpha messenger RNA (mRNA) in Hep3B cells. The maximal increase was induced by 2% isoflurane, and the cells incubated with 2% isoflurane for 4-8 h expressed the highest protein. Similarly, HIF-1alpha transcriptional activity was higher in Hep3B cells exposed to 2% isoflurane for 16 h than that in control cells. The combination of 2% isoflurane and desferrioxamine, a hypoxia mimetic, caused a higher level of HIF-1alpha protein than that induced by 2% isoflurane alone. Reoxygenation and inhibitor of proteasome pathway MG132 did not affect the isoflurane-induced HIF-1alpha protein accumulation. Cycloheximide, an inhibitor for protein synthesis, completely abrogated the induction of HIF-1alpha protein by isoflurane. Isoflurane stimulated heme oxygenase 1, inducible nitric oxide synthase, and VEGF mRNA expression in a concentration-dependent manner, and inactivation of HIF-1alpha attenuated the induction of VEGF mRNA by isoflurane.


CONCLUSIONS
Isoflurane can up-regulate HIF-1alpha and enhance HIF-1-responsive genes heme oxygenase 1, inducible nitric oxide synthase, and VEGF mRNA expression in Hep3B cells. The induction of HIF-1alpha by isoflurane does not involve protein degradation but depends on translation pathway.
Li
Qi Fang
QF
Department of Anesthesiology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, 1630 Dongfang Road, Shanghai 200127, China.
Wang
Xiang Rui
XR
Yang
Yue Wu
YW
Su
Dian San
DS
D000199:Actins / Q000096:biosynthesis; D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis*; D007530:Isoflurane / Q000494:pharmacology*; D052247:Nitric Oxide Synthase Type II / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis
D000199:Actins; D018685:Anesthetics, Inhalation; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D034741:RNA, Small Interfering; D042461:Vascular Endothelial Growth Factor A; D007530:Isoflurane; D052247:Nitric Oxide Synthase Type II