Desflurane preconditioning inhibits endothelial nuclear factor-kappa-B activation by targeting the proximal end of tumor necrosis factor-alpha signaling.
BACKGROUND
Volatile anesthetics interfere with inflammatory cytokine production and expression of adhesion molecules which are critical for ischemia reperfusion induced injury. Nuclear factor (NF)-kappaB has been reported to be suppressed in this process, but the detailed molecular mechanism is still unclear.


METHODS
In this study, ECV304 (a human umbilical vein endothelial cell line) was preconditioned with 30 min desflurane (1 minimal alveolar concentration), after 15 min washout, 30 min anoxia, and 60 min reoxygenation was performed. ECV304 was finally stimulated with tumor necrosis factor (TNF)-alpha (10 ng/mL). Control groups, which were not preconditioned and/or not stimulated, were also included in the protocol. IkappaB-alpha, phospho-IkappaB-alpha, phospho-IkappaB kinase (IKKalpha)/IKKbeta, and phospho-p38 were detected by Western blotting. The nuclear NF-kappaB p65 subunit was measured by subcellular fractionation and Western blotting. The surface expression of TNF-R1 was measured by flow cytometry. Receptor-associated signaling adaptors, e.g., TNF receptor-associated factor 2 (TRAF2) and IKK-alpha, were evaluated by immunoprecipitation by TNF-R1 antibody and subsequent Western blotting.


RESULTS
Desflurane preconditioning inhibits IkappaB-alpha phosphorylation, degradation, and p65 nuclear localization. Desflurane also affects p38 phosphorylation, which is needed for optimal inflammatory response. The phosphorylation of IKKalpha/IKKbeta was suppressed by preconditioning while the surface abundance of TNF-R1 was not affected. The association of TRAF2 and IKK-alpha with TNF-R1 was compromised by desflurane.


CONCLUSIONS
Our results suggest that the molecular target of desflurane in the NF-kappaB pathway is upstream of IKK activation. The abundance of TNF-R1 on the cell membrane is not affected by anesthetic preconditioning. We suggest that desflurane preconditioning targets the proximal end of TNF-alpha signaling.
Li
Yuan
Y
Department of Anesthesiology, Zhongshan Hospital, Fudan University, Fenglin Road 180, Shanghai 200032, China.
Zhang
Xiaonan
X
Zhu
Biao
B
Xue
Zhanggang
Z
D021581:Active Transport, Cell Nucleus; D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D002460:Cell Line; D000077335:Desflurane; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D020672:I-kappa B Proteins / Q000378:metabolism; D007530:Isoflurane / Q000031:analogs & derivatives* / Q000494:pharmacology; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015427:Reperfusion Injury / Q000378:metabolism* / Q000517:prevention & control; D015398:Signal Transduction / Q000187:drug effects*; D047992:TNF Receptor-Associated Factor 2 / Q000378:metabolism; D051996:Transcription Factor RelA / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D018685:Anesthetics, Inhalation; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D047888:Receptors, Tumor Necrosis Factor, Type I; D011994:Recombinant Proteins; D047992:TNF Receptor-Associated Factor 2; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D000077335:Desflurane; D007530:Isoflurane; D051550:I-kappa B Kinase; D048051:p38 Mitogen-Activated Protein Kinases