title: Cytoprotective effects of the volatile anesthetic sevoflurane are highly dependent on timing and duration of sevoflurane conditioning: findings from a human, in-vitro hypoxia model.
abstract: Using animal models, volatile anesthetics have been recognized for their neuroprotective effects. Nevertheless, there is still disagreement about the optimal duration and timing of conditioning with the volatile anesthetic sevoflurane in the human system. In the study presented, we employed a human neuronal cell culture model to investigate the effects of hypoxia and to evaluate potential cytoprotective properties of different sevoflurane conditioning strategies. Sevoflurane was applied to human IMR-32 cells in which hypoxic conditions were induced for 2h using our recently described two-enzyme model (Zitta et al., Eur. J. Pharmacol., 2010). Cellular effects of hypoxia and sevoflurane conditioning were evaluated by lactate dehydrogenase (LDH) measurements, brightfield microscopy, ELISAs, cytometric bead arrays, Westernblotting and RT-PCR. Hypoxia increased the release of LDH into the culture medium after 24h (normoxia: 0.15+/-0.02 a.u; hypoxia: 0.69+/-0.08 a.u, P<0.001) and expression of hypoxia associated genes HIF-1alpha, VEGF, catalase. Cytoprotective effects were observed in cultures that received sevoflurane for 30 min before hypoxia (preconditioning: 0.41+/-0.07 a.u., P<0.01) and for 30 min during the hypoxic period (intraconditioning: 0.20+/-0.01 a.u., P<0.001). Application of sevoflurane after the hypoxic insult did not lead to cytoprotection (postconditioning: 0.73+/-0.12a.u., P>0.05). Conditioning with sevoflurane for a total of 3h before, during and after hypoxia, however, resulted in an enhanced release of LDH (periconditioning: 0.97+/-0.10a.u., P<0.01) and additional cell damage. Hypoxia and sevoflurane intraconditioning were associated with changes in erk1/2 phosphorylation (T202/Y204) and HIF-1alpha protein levels, whereas phosphorylation of akt (S473) was not significantly altered. Our results suggest short pre- and intraconditioning with sevoflurane as most potent strategies to reduce hypoxia induced neuronal cell damage.
authors.affiliation: Department of Anesthesiology and Intensive Care Medicine, University Hospital Schleswig-Holstein, Campus Kiel, Kiel, Germany.
authors.forename: Karina
authors.initials: K
authors.lastname: Zitta
authors.forename: Patrick
authors.initials: P
authors.lastname: Meybohm
authors.forename: Berthold
authors.initials: B
authors.lastname: Bein
authors.forename: Henning
authors.initials: H
authors.lastname: Ohnesorge
authors.forename: Markus
authors.initials: M
authors.lastname: Steinfath
authors.forename: Jens
authors.initials: J
authors.lastname: Scholz
authors.forename: Martin
authors.initials: M
authors.lastname: Albrecht
mesh_terms: D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D002374:Catalase / Q000378:metabolism; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D019610:Cytoprotection; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008738:Methyl Ethers / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010766:Phosphorylation; D000077149:Sevoflurane; D013997:Time Factors; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D018685:Anesthetics, Inhalation; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008738:Methyl Ethers; D018696:Neuroprotective Agents; D042461:Vascular Endothelial Growth Factor A; D000077149:Sevoflurane; D007770:L-Lactate Dehydrogenase; D002374:Catalase; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3