Hyperglycemia Adversely Modulates Endothelial Nitric Oxide Synthase during Anesthetic Preconditioning through Tetrahydrobiopterin– and Heat Shock Protein 90–mediated Mechanisms

Abstract

Background Endothelial nitric oxide synthase activity is regulated by tetrahydrobiopterin (BH4) and heat shock protein 90. We tested the hypothesis that hyperglycemia abolishes anesthetic preconditioning (APC) through BH4- and heat shock protein 90-dependent pathways. Methods Myocardial infarct size was measured in rabbits in the absence or presence of APC (30 min of isoflurane), with or without hyperglycemia, and in the presence or absence of the BH4 precursor sepiapterin. Isoflurane-dependent nitric oxide production was measured (ozone chemiluminescence) in human coronary artery endothelial cells cultured in normal (5.5 mM) or high (20 mM) glucose conditions, with or without sepiapterin (10 or 100 µM). Results APC decreased myocardial infarct size compared with control experiments (26±6 vs 46±3%, respectively; P<0.05), and this action was blocked by hyperglycemia (43±4%). Sepiapterin alone had no effect on infarct size (46±3%) but restored APC during hyperglycemia (21±3%). The beneficial actions of sepiapterin to restore APC were blocked by the nitric oxide synthase inhibitior N (G)-nitro-L-arginine methyl ester (47±2%) and the BH4 synthesis inhibitor N-acetylserotonin (46±3%). Isoflurane increased nitric oxide production to 177±13% of baseline, and this action was attenuated by high glucose concentrations (125±6%). Isoflurane increased, whereas high glucose attenuated, intracellular BH4/BH2 (high-performance liquid chromatography), heat shock protein 90-endothelial nitric oxide synthase co-localization (confocal microscopy), and endothelial nitric oxide synthase activation (immunoblotting). Sepiapterin increased BH4/BH2 and dose-dependently restored nitric oxide production during hyperglycemic conditions (149±12 and 175±9%; 10 and 100 µM, respectively). Conclusion The results indicate that tetrahydrobiopterin and heat shock protein 90-regulated endothelial nitric oxide synthase activity play a central role in cardioprotection that is favorably modulated by volatile anesthetics and dysregulated by hyperglycemia. Enhancing the production of BH4 may represent a potential therapeutic strategy.

Decreased tetrahydrobiopterin and disrupted association of Hsp90 with eNOS by hyperglycemia impair myocardial ischemic preconditioning

“…Myocardial BH4 was quantified by HPLC with an electrochemical detector (ESA Biosciences CoulArray Sys- tem, Chelmsford, MA), as previously described (1). Briefly, rabbit myocardium from the risk region was homogenized in 50 mM phosphate buffer (pH 2.6) containing 0.2 mM diethylenetriaminepentaacetic acid and 1 mM dithioerytrithol.…”

“…Nitrite concentration corresponding to the stable byproduct of NO released by HCAECs in aqueous solution was quantified by ozone chemiluminescence (1). Samples (20 l) were refluxed in glacial acetic acid containing potassium iodide and nitrite quantified in a NO chemiluminescence analyzer (Sievers Instruments, Boulder, CO).…”

“…Biochemical and pharmacological evidence indicates that BH 4 deficiency or disruption of the association of Hsp90 with eNOS results in "uncoupling" of eNOS with L-arginine, leading to the production of superoxide anion rather than NO by this enzyme (4,21,41,44,51). Interestingly, diabetes and HG have been shown to modulate these regulators of eNOS activity (1). For example, overproduction of reactive oxygen species by mitochondria during diabetes and HG results in oxidation of intracellular BH 4 to the catalytically incompetent species dihydrobiopterin.…”

A Recipe for Perioperative Cardioprotection

“…and "Does anesthetic choice matter?" Volatile anesthetics have been shown to precondition and postcondition myocardium through a variety of signaling molecules and proteins, 6 including endothelial nitric oxide synthase, heat shock protein 90, tetrahydrobiopterin, mitochondrial and sarcolemmal adenosine triphosphate-regulated potassium channels, intracellular survival kinases, and membrane-bound receptors. The timing and mode of administration (stuttered versus continuous), 7 as well as the dose of volatile anesthetic, influences the efficacy of these drugs to reduce ischemic injury.…”

Glycemic Control in Cardiac Surgery

“…Others effects of excessive glucose levels include the impairment in hepatic functions, abolishment of ischemic preconditioning, and protein glycosylation (56). Cardiac Surgery has been traditionally considered a highly-invasive procedure (57, 58) and taking in account that the hyperglycemic response is increased by anesthetics and the preoperative emotional stress (59), this major surgery, induces the release of several counterregulatory hormones and deeply modifies the metabolism of carbohydrates, causing increased hepatic gluconeogenesis, insulin resistance in various peripheral tissues, and the relative insulin production deficiency (60,61).…”