Hemorrhagic shock and nitric oxide release from erythrocytic nitric oxide synthase: A quantitative analysis

Department of Biomedical Engineering, School of Medicine, Johns Hopkins University, 613 Traylor Building, 720 Rutland Avenue, Baltimore, MD 21205, USA.
Microvascular Research (Impact Factor: 2.13). 04/2009; 78(1):107-18. DOI: 10.1016/j.mvr.2009.02.009
Source: PubMed


A large loss of blood during hemorrhage can result in profound shock, a state of hypotension associated with hemodynamic abnormalities. One of the hypotheses to account for this collapse of homeostasis is that the production of nitric oxide (NO), a gas molecule that dilates blood vessels, is significantly impaired during hemorrhage, resulting in a mismatch between O(2) delivery and the metabolic activity in the tissues. NO can be released from multiple sources in the vasculature. Recent studies have shown that erythrocytes express functional endothelial nitric oxide synthase (NOS3), which potentially serves as an intraluminal NO source. NO delivery from this source is complex: erythrocytes are not only NO producers but also act as potent sinks because of the high affinity of NO for hemoglobin. To test our hypothesis that the loss of erythrocytic NOS3 during hemorrhage contributes to NO deficiency-related shock, we have constructed a multicellular computational model that simulates NO production and transport to allow us to quantify the loss of NO under different hemorrhagic conditions. Our model shows that: (1) during mild hemorrhage and subsequent hemodilution (hematocrit >30%), NO from this intraluminal source is only slightly decreased in the vascular smooth muscle, but the NO level is significantly reduced under severe hemorrhagic conditions (hematocrit <30%); (2) whether a significant amount of NO from this source can be delivered to vascular smooth muscle is strongly dependent on the existence of a protective mechanism for NO delivery; (3) if the expression level of NOS3 on erythrocytes is similar to that on endothelial cells, we estimate approximately 13 pM NO at the vascular smooth muscle from this source when such a protective mechanism is involved. This study provides a basis for detailed studies to characterize the impairment of NO release pathways during hemorrhage and yield important insights for the development of resuscitation methods.

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Available from: Aleksander S Popel
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    • "In addition, RBC-NOS resembles endothelium-derived eNOS in that it is specifically stimulated by the substrate larginine , it is sensitive to common NOS inhibitors, and its activity depends on the intracellular calcium level (Fulton et al., 2001). Recently, computational modeling (Chen et al., 2009) simulated NO production from eNOS in erythrocytes and its transport through an arteriole. It was found that the expression level of eNOS in erythrocytes is similar to that in endothelium, and researchers can predict a level NO in the vascular smooth muscle from this RBCs source. "
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    DESCRIPTION: new simple method for measuring endothelial NOS in clinical researches
    Full-text · Research · Jan 2016
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    • "It has been argued that RBC have a dual role in tissue oxygenation: in addition to their wellknown oxygen carrier function, RBC may contribute to the regulation of local blood flow with NO playing a central role in this process [35]. NO synthesized by RBC NOS activity has been hypothesized to contribute to the RBC-originated NO pool, although Chen, et al. reported that the amount of NO generated by RBC NOS may not represent a physiologically important fraction of total NO bioavailability at vascular wall [36]. Experimental studies directly exploring the contribution of the enzymatic generation of NO by RBC to local vasomotor control do not yet exist. "
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    ABSTRACT: Red blood cells (RBC) play an important role in the balance between generation and scavenging of nitric oxide (NO) and hence its local bioavailability and influence on vasomotor control. Previous studies have reported increased NO levels in RBC suspensions subsequent to exposure to shear forces; the present study was designed to further investigate changes in intracellular NO concentration and possible mechanisms involved for RBC exposed to well-controlled shear forces. Attached human RBC were subjected to shear stresses up to 0.1Pa in a parallel-plate flow channel; fluorescent methods were used to monitor changes in intracellular NO and calcium concentrations. Intracellular NO concentration, estimated by the fluorescence level of 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM), increased sharply within 30s following the application of shear stress between 0.013 and 0.1Pa. This increase was only partially prevented by the absence of l-arginine and by the presence of l-N-acetyl-methyl-arginine (L-NAME), strongly suggesting that this response was in part related to the activation of NO-synthase (NOS) enzyme. The increase in intracellular NO concentration under shear stress was also inhibited by calcium chelation in the suspending medium, indicating the role of calcium entry for NOS activation. Increases of intracellular calcium concentrations under the same shearing conditions were demonstrated by monitoring Fluo-3/AM fluorescence in RBC exposed to shear stress. Serine 1177 phosphorylated NOS protein, the activated form of the enzyme determined by immunohistochemistry, was found to be significantly increased following the exposure of RBC to 0.1Pa shear stress for 1min. These data confirm that RBC possess a NOS enzyme that is actively synthesizing NO and activated by effective shear forces. The data also suggest that there may be additional (e.g., non-enzymatic) NO generating mechanisms in RBC that are also enhanced under shear stress.
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    • "Computer simulations indicate that, as has been shown for other intraerythrocytic pathways, whether a significant amount of NO from this source can be delivered to vascular smooth muscle is strongly dependent on the existence of a protected mechanism for NO delivery. If the expression level of NOS3 in erythrocytes is similar to that in endothelial cells, a combined NOS biochemical pathway analysis model [16] [17] and NO transport model [13] would predict a level of ∼13 pM NO in the vascular smooth muscle from this source, given the existence of such a protected NO transport mechanism [15] "
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    ABSTRACT: Nitric oxide (NO) is a potent regulator of vascular tone and hemorheology. The signaling function of NO was largely unappreciated until approximately 30 years ago, when the endothelium-derived relaxing factor (EDRF) was identified as NO. Since then, NO from the endothelium has been considered the major source of NO in the vasculature and a contributor to the paracrine regulation of blood hemodynamics. Because NO is highly reactive, and its half-life in vivo is only a few seconds (even less in the bloodstream), any NO bioactivity derived from the intraluminal region has traditionally been considered insignificant. However, the availability and significance of NO signaling molecules derived from intraluminal sources, particularly erythrocytes, have gained attention in recent years. Multiple potential sources of NO bioactivity have been identified in the blood, but unresolved questions remain concerning these proposed sources and how the NO released via these pathways actually interacts with intravascular and extravascular targets. Here we review the hypotheses that have been put forward concerning blood-borne NO and its contribution to hemorheological properties and the regulation of vascular tone, with an emphasis on the quantitative aspects of these processes.
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