title: Hypoxic potentiation of nitrite effects in human vessels and platelets.
abstract: Previous studies in non-human blood vessels and in platelets have demonstrated that under hypoxic conditions release of NO from nitrite (NO2(-)) is potentiated by deoxyhaemoglobin. In the current study, we characterized hypoxic potentiation of NO2(-) effects in human vasculature and platelets in vitro, addressing underlying mechanisms. The vasodilator efficacy of NO2(-), in comparison with glyceryl trinitrate (GTN), was evaluated in vitro, using segments of human saphenous vein. Under hypoxic conditions, there was a leftward shift of the NO2(-) concentration-response curve (EC50: 22 μM in hyperoxia vs 3.5 μM in hypoxia; p<0.01), but no significant potentiation of GTN effect. In the presence of red blood cells, hypoxic potentiation of NO2(-) vasodilator effect was accentuated. In whole blood samples and platelet-rich plasma (PRP) we assessed inhibition of platelet aggregation by NO2(-) (1mM), in comparison with that of sodium nitroprusside (SNP, 10 μM). In individual subjects (n=37), there was a strong correlation (r=0.75, p<0.0001) between anti-aggregatory effects of NO2(-) and SNP in whole blood, signifying that resultant sGC activation underlies biological effect and responses to NO2(-) are diminished in the presence of NO resistance. In PRP, the effects of NO2(-) were less pronounced than in whole blood (p=0.0001), suggesting an important role of Hb (within RBCs) in the bioconversion of NO2(-) to NO. Inhibition of platelet aggregation by NO2(-) was almost 3-fold greater in venous than in arterial blood (p<0.0001), and deoxyHb concentration directly correlated (r=0.69, p=0.013) with anti-aggregatory response. Incremental hypoxia applied to venous blood samples (in hypoxic chamber) caused a progressive increase in both deoxyHb level and anti-aggregatory effect of NO2(-). When subjects inhaled a 12% O2 mixture for 20 min, there was a 3-fold rise in blood deoxyHb fraction (p<0.01). In PRP, response to NO2(-) also increased under hypoxia, and was further enhanced (p<0.01) by deoxyHb. Furthermore, deoxyHb exerted significant anti-aggregatory effects even in the absence of added NO2(-), suggesting a role for endogenous NO2(-). The results of this work provide further mechanistic insights into hypoxic potentiation of vasodilator and anti-aggregatory actions of NO2(-). In human saphenous veins and blood, the balance of evidence suggests differential rates of NO release from NO2(-) (largely modulated by deoxyHb) as the fundamental mechanism.
authors.affiliation: Cardiology Unit, Basil Hetzel Institute, The Queen Elizabeth Hospital, University of Adelaide, Woodville, Australia. Electronic address: rustem.dautov@adelaide.edu.au.
authors.forename: Rustem F
authors.initials: RF
authors.lastname: Dautov
authors.affiliation: Cardiology Unit, Basil Hetzel Institute, The Queen Elizabeth Hospital, University of Adelaide, Woodville, Australia.
authors.forename: Irene
authors.initials: I
authors.lastname: Stafford
authors.affiliation: Cardiology Unit, Basil Hetzel Institute, The Queen Elizabeth Hospital, University of Adelaide, Woodville, Australia.
authors.forename: Saifei
authors.initials: S
authors.lastname: Liu
authors.affiliation: Cardiac and Thoracic Surgical Unit, Flinders Medical Center, Flinders University, Bedford Park, Australia.
authors.forename: Hugh
authors.initials: H
authors.lastname: Cullen
authors.affiliation: Centre for Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, B15 2TT, UK.
authors.forename: M
authors.initials: M
authors.lastname: Madhani
authors.affiliation: Cardiology Unit, Basil Hetzel Institute, The Queen Elizabeth Hospital, University of Adelaide, Woodville, Australia.
authors.forename: Yuliy Y
authors.initials: YY
authors.lastname: Chirkov
authors.affiliation: Cardiology Unit, Basil Hetzel Institute, The Queen Elizabeth Hospital, University of Adelaide, Woodville, Australia.
authors.forename: John D
authors.initials: JD
authors.lastname: Horowitz
mesh_terms: D000328:Adult; D000368:Aged; D001792:Blood Platelets / Q000187:drug effects*; D001808:Blood Vessels / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D018496:Hyperoxia; D008297:Male; D009573:Nitrites / Q000494:pharmacology*; D010974:Platelet Aggregation / Q000187:drug effects*; D013329:Structure-Activity Relationship
chemical_list: D009573:Nitrites
keywords: Haemoglobin; Hypoxia; Nitric oxide resistance; Nitrite; Platelet aggregation; Vasodilation