Beta2-adrenergic agonist protects human endothelial cells from hypoxia/reoxygenation injury in vitro.
OBJECTIVE
Circulatory shock results in hypoxia/reoxygenation processes that lead to the release of reactive oxygen species, endothelial injury, and multiple organ failure. Previous data suggest that beta2-adrenergic agonists prevent endothelial dysfunction. The study aimed at determining whether the beta2-adrenergic agonist formoterol protects endothelial cells against hypoxia/reoxygenation injury in vitro.


METHODS
Prospective controlled trial.


METHODS
University hospital research laboratory.


METHODS
Cultured human umbilical vein endothelial cells (HUVECs).


METHODS
Confluent HUVECs were sealed in a flow-through chamber mounted on an inverted microscope and perfused with a constant flow of Krebs medium. After 1 hr of equilibration, HUVECs underwent 2 hrs of hypoxia and 1 hr of reoxygenation. Cell death at the end of reoxygenation and reactive oxygen species formation were assessed with fluorescent probes propidium iodide and 2',7'-dichlorodihydrofluorescein diacetate, respectively. The effects of the beta2-adrenergic agonist formoterol, the beta2-adrenergic antagonist ICI 118,551 and the nitric oxide synthase inhibitor L-NNA were investigated. Statistical analysis was performed with analysis of variance followed by post hoc Fisher's test.


RESULTS
Hypoxia/reoxygenation increased cell death (hypoxia/reoxygenation 29 +/- 4% vs. control 1 +/- 5%, p < .05) and endothelial reactive oxygen species production (hypoxia/reoxygenation 126 +/- 4% vs. control 108 +/- 4%, p < .05). Formoterol reduced cell death in a concentration-dependent manner (EC95 = 10 mol/L) and reduced endothelial reactive oxygen species production (hypoxia/reoxygenation + formoterol EC95 109 +/- 4% vs. hypoxia/reoxygenation 126 +/- 4%, p < .05). When added to formoterol EC95, ICI 118,551 and L-NNA abolished the formoterol-induced cell protection and reduced reactive oxygen species production.


CONCLUSIONS
These results indicate that formoterol reduces endothelial cell death and reactive oxygen species production in this in vitro hypoxia/reoxygenation model. These effects are beta2-adrenergic specific and are partially mediated by nitric oxide synthase.
Pottecher
Julien
J
Department of Anesthesiology and Critical Care, University Hospital, Le Kremlin Bicêtre, France.
Cheisson
Gaëlle
G
Huet
Olivier
O
Laplace
Christian
C
Vicaut
Eric
E
Mazoit
Jean X
JX
Benhamou
Dan
D
Duranteau
Jacques
J
D000318:Adrenergic beta-Agonists / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D015687:Cell Hypoxia / Q000187:drug effects* / Q000502:physiology; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects*; D004983:Ethanolamines / Q000494:pharmacology*; D000068759:Formoterol Fumarate; D006801:Humans; D066298:In Vitro Techniques; D019001:Nitric Oxide Synthase / Q000032:analysis / Q000378:metabolism; D010477:Perfusion; D017382:Reactive Oxygen Species / Q000378:metabolism; D012016:Reference Values; D015427:Reperfusion Injury / Q000517:prevention & control*; D012680:Sensitivity and Specificity
D000318:Adrenergic beta-Agonists; D004983:Ethanolamines; D017382:Reactive Oxygen Species; D019001:Nitric Oxide Synthase; D000068759:Formoterol Fumarate