Received January 28, 2005
Revised February 24, 2005
Accepted after revision March 4, 2005

¹ Fundación Jiménez Díaz
² Hospital Clinico San Carlos
³ Hospital Clínico San Carlos
⁴ Fundación Jimenez Díaz

^* To whom correspondence should be addressed. E-mail: lcarinv.hcsc{at}salud.madrid.org.

	Abstract

Our aim was to analyze endothelial hypoxic preconditioning after hypoxia-reperfusion (HR). Endothelial functionality was analyzed through the vasorelaxation responses to acetylcholine (Ach) and the level of serine¹¹⁷⁷ phosphorylated endothelial nitric oxide synthase (eNOS) (ser¹¹⁷⁷-eNOS) measured by Western blot in vitro hypoxic preconditioned (P+HR) isolated rat aortic segments. Relaxation to Ach was reduced in phenylephrine-precontracted aortic segments after HR (Control: IC₅₀=5x10^-8 ± 2.5 mol/l; HR: IC₅₀=3x10^-7 ± 1.2 mol/l; p<0.05). Ach-dependent vasodilatation was improved by P+HR. The content of ser¹¹⁷⁷-eNOS in the HR segments was 1.5 fold lower than in P+HR. Confocal microscopy showed an increased content of both superoxide anion and peroxynitrite in the vascular wall of HR aortic segments, which it was reduced by P+HR. Geldanamycin (10 µg/mL), an agent known to inhibit heat shock protein 90 (hsp90), reduced the level of ser¹¹⁷⁷-eNOS in P+HR aortic segments. However, in the presence of geldanamycin endothelial hypoxic preconditioning persisted. As conclusions, short periods of hypoxia induced endothelial hypoxic preconditioning that was accompanied by enhanced levels of ser¹¹⁷⁷-eNOS in the vascular wall. The fact that endothelial hypoxic preconditioning persisted in the presence of geldanamycin suggests that other molecular mechanisms are involved in the endothelial adaptation to HR injury. Keywords: endothelium, hypoxia, hypoxic preconditioning, endothelial nitric oxide synthase, superoxide anion.

Key Words: Anion, Endothelium, Hypoxia