Case Reports  |   September 2006
Inhaled Nitric Oxide for Treatment of Sickle Cell Stroke
Author Affiliations & Notes
  • Pedro Montero-Huerta, M.D.
  • Dean R. Hess, Ph.D.
  • C Alvin Head, M.D.
  • * Instructor, ‡ Professor and Chairman, Anesthesiology and Perioperative Medicine, Medical College of Georgia, Augusta, Georgia. † Associate Professor, Harvard Medical School, Massachusetts General Hospital, Boston, Massachusetts.
Article Information
Case Reports / Central and Peripheral Nervous Systems
Case Reports   |   September 2006
Inhaled Nitric Oxide for Treatment of Sickle Cell Stroke
Anesthesiology 9 2006, Vol.105, 619-621. doi:
Anesthesiology 9 2006, Vol.105, 619-621. doi:
STROKE is a highly fatal complication of sickle cell disease (SCD) in children, particularly between the ages of 4 and 15 yr. Children with SCD carry a 300-fold increased risk for stroke; consequently, sickle cell anemia is the most common cause of childhood stroke.1 We report the case of a 13-yr-old African-American boy diagnosed at age 6 months old with hemoglobin SS disease, who had a nonhemorrhagic stroke after a routine anesthetic and was treated with inhaled nitric oxide (INO).
Case Report
This child underwent a simple, uneventful surgical procedure for central venous access to allow for exchange transfusion to achieve a hemoglobin S value of less than 20%. He received general anesthesia with sevoflurane, oxygen, midazolam, and fentanyl. There was no hypotension or hypoxia during the procedure. Three days before this surgical procedure, he received an erythrocyte transfusion. His preoperative laboratory results were as follows: erythrocyte count, 4.99 × 106/mm3; hemoglobin, 15.1 g/dl; hematocrit, 44.1%; mean corpuscular volume, 84 mm3; erythrocyte distribution width, 14.2%; leukocytes, 12.5 × 103/mm3; neutrophils, 62%; lymphocytes, 28%; monocytes, 3%; eosinophils, 4%; basophils, 1%.
Shortly after surgery in the postoperative care unit, his mother noted that he became unresponsive preceding seizure activity. He was emergently taken for magnetic resonance imaging and magnetic resonance angiography, which revealed right-sided cerebrovascular accident in the distribution of the right middle cerebral artery, and severe bilateral internal carotid artery stenosis. This stroke limited left-sided facial, arm, and leg movements. He was transferred to the Pediatric Intensive Care Unit and, after consultation with the Neurology and Stroke Service, it was decided to treat him with an exchange transfusion. Anesthesiology was also consulted to review the surgical record, and after concluding that the anesthetic and surgical procedures were within the normal standards of care, a trial of INO for this patient was discussed with the parent.
United States Food and Drug Administration approval for off-label use of INO was obtained; emergency institutional review board approval was granted (Partners Human Research Committee, Boston, Massachusetts), and the patient’s mother signed a consent-to-treat form before the exchange transfusion. INO was administered by facemask at 80 parts per million by volume. Blood samples were obtained before INO and after 3 and 22 h of therapy.
To estimate endogenous nitric oxide, we measured plasma nitrite and nitrate levels, which are stable end products of nitric oxide. Indirect measurements of nitric oxide are needed because its half-life in whole blood is in milliseconds.2 
Plasma was separated from whole blood by centrifugation, and measurements of plasma nitric oxide metabolites (NOx) were made using a Sievers Nitric Oxide Analyzer 280 (Sievers Instruments, Inc., Boulder, CO). Samples (5 μl) were injected into the headspace through the septum of an oxygen-free purger vessel containing 5–6 ml vanadium (III) chloride in HCl. The vanadium III, heated to 94°C by a circulating water bath, reduced nitrites and nitrates to nitric oxide gas, which was measured by the NOA 280 chemiluminescence detector (Sievers Instruments, Inc.). NOx concentrations were determined as area under the curve of three separate injections of samples and integrating the peaks. The areas were compared with calibration curves produced by the injection of sodium nitrate standards.
Nitric oxide metabolite levels were dramatically low (14.8 μm) before initiating INO therapy (fig. 1). NOx levels increased (82 μm) after 3 h of nitric oxide breathing and were further increased (106 μm) at 22 h (fig. 1). The patient became neurologically responsive within 3 h of INO therapy and before exchange transfusion. INO was continued for 48 h, at which time the patient had a near complete neurologic recovery.
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx). 
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx). 
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx). 
Sickle cell disease  is a genetic disorder whose manifestations are caused by a single point mutation that results in the substitution of valine for glutamic acid at the sixth position β-globin subunit.3 Sickle hemoglobin forms polymers during deoxygenation. When deoxygenated, sickle hemoglobin aggregates and produces a viscous gel composed of multistranded helical polymers, resulting in rigid and deformed erythrocytes. In the microvasculature, adhesion of the sickle erythrocytes to the vascular endothelium occurs. This produces slowing and obstruction of the microcirculation, creating localized ischemia and infarction. The resulting acute and chronic organ damage is a major cause of pain, morbidity, and mortality associated with SCD.4 By age 20 yr, approximately 11% of homozygous SCD patients will experience a stroke.1,5,6 Most strokes in SCD pediatric patients are nonhemorragic.1 It is unclear whether general anesthesia or surgical trauma triggered the acute stroke in this child. However, the close temporal relation with the minor surgical procedure suggests this to be the case.
Available evidence supports the occurrence of ischemia-reperfusion injury–like events in the vasculature of SCD patients due to erythrocyte adhesion. This injury produces a dysfunctional endothelium favoring a procoagulant state.7 Moreover, such dysfunctional endothelium is less capable of producing nitric oxide.8–10 In addition, endogenous nitric oxide is avidly scavenged and consumed by the large amounts of free hemoglobin and by the overproduction of reactive oxygen species occurring in SCD.11–13 These factors acting together may significantly reduce nitric oxide bioavailability and could play an important role in the pathogenesis of SCD stroke. This may explain the low NOx measured in this patient before INO therapy. It is well known that nitric oxide is a central player regulating platelet aggregation, cell adhesion, and vascular tone. Therefore, repletion of nitric oxide by inhalation may provide benefit in this condition.
Inhaled nitric oxide could be beneficial in the treatment of stroke in sickle cell in a multifactorial way. First, it increases nitric oxide bioavailability. Second, it improves blood flow and oxygenation as a result of preventing erythrocyte, platelet, and leukocyte adhesion to the vascular endothelium. Third, as we previously demonstrated using a mouse model of SCD, INO has protective properties in hypoxic stress.14 Although debated, this protective effect may be related to an increase in oxygen affinity created by a reduction in sickle hemoglobin polymers.15,16 
Although plasma NOx levels were not measured before surgery, it is possible that this patient may have had a subclinical inflammatory process ongoing, allowing him to be at increased risk for stroke during his anesthesia and surgery.
In this case, plasma NOx levels were extremely low, which correlates with NOx levels published by others during SCD crisis.17,18 INO therapy was associated with clinical improvement in this child before conventional therapy, including blood exchange transfusion. The rapid improvement in neurologic status was dramatic, suggesting a relation to INO therapy. The clinical improvement was also associated with an increase in plasma NOx (fig. 1). After 24 h of nitric oxide breathing, magnetic resonance imaging and magnetic resonance angiography analysis did not show significant changes. However, conventional magnetic resonance imaging alone has been reported to correlate poorly with physical recovery in stroke related to SCD.19 A combination of different radiologic techniques, such as diffusion and perfusion-weighted magnetic resonance imaging preferably in conjunction with positron emission tomography, may have better assessed the effects of nitric oxide in poststroke recovery.19–21 Although recovery from sickle stroke is possible without INO therapy, our hematologist, who performed the exchange transfusion, believed this stroke would not have resolved naturally. In addition, reduced plasma NOx suggests reduced nitric oxide bioavailability. Hence, in this patient, preoperative NOx measurements may have proven helpful in identifying a risk factor for operative complications. However, without proof, these observations should be interpreted with caution, and clinical trials should be considered to evaluate the potential role of INO in the treatment of sickle stroke.
The authors thank David H. Ebb, M.D. (Clinical Director; Pediatric Brain Tumor Program, MassGeneral CancerCare for Children; Assistant Pediatrician, Massachusetts General Hospital; Assistant Professor of Pediatrics, Harvard Medical School, Boston, Massachusetts), who was the patient’s attending physician.
Ohene-Frempong K, Weiner SJ, Sleeper LA, Miller ST, Embury S, Moohr JW, Wethers DL, Pegelow CH, Gill FM: Cerebrovascular accidents in sickle cell disease: Rates and risk factors. Blood 1998; 91:288–94Ohene-Frempong, K Weiner, SJ Sleeper, LA Miller, ST Embury, S Moohr, JW Wethers, DL Pegelow, CH Gill, FM
Liu X, Miller MJ, Joshi MS, Sadowska-Krowicka H, Clark DA, Lancaster JR Jr: Diffusion-limited reaction of free nitric oxide with erythrocytes. J Biol Chem 1998; 273:18709–13Liu, X Miller, MJ Joshi, MS Sadowska-Krowicka, H Clark, DA Lancaster, JR
Pauling L, Itano HA, Singer SJ, Wells IC: Sickle cell anemia, a molecular disease. Science 1949; 110:543–8Pauling, L Itano, HA Singer, SJ Wells, IC
Horne MK III: Sickle cell anemia as a rheologic disease. Am J Med 1981; 70:288–98Horne, MK
Adams RJ, Nichols FT III, Aaslid R, McKie VC, McKie K, Carl E, Stephens S, Thompson WO, Milner P, Figueroa R: Cerebral vessel stenosis in sickle cell disease: Criteria for detection by transcranial Doppler. Am J Pediatr Hematol Oncol 1990; 12:277–82Adams, RJ Nichols, FT Aaslid, R McKie, VC McKie, K Carl, E Stephens, S Thompson, WO Milner, P Figueroa, R
McCarville MB, Li C, Xiong X, Wang W: Comparison of transcranial Doppler sonography with and without imaging in the evaluation of children with sickle cell anemia. AJR Am J Roentgenol 2004; 183:1117–22McCarville, MB Li, C Xiong, X Wang, W
Hebbel RP, Osarogiagbon R, Kaul D: The endothelial biology of sickle cell disease: Inflammation and a chronic vasculopathy. Microcirculation 2004; 11:129–51Hebbel, RP Osarogiagbon, R Kaul, D
French JA II, Kenny D, Scott JP, Hoffmann RG, Wood JD, Hudetz AG, Hillery CA: Mechanisms of stroke in sickle cell disease: Sickle erythrocytes decrease cerebral blood flow in rats after nitric oxide synthase inhibition. Blood 1997; 89:4591–9French, JA Kenny, D Scott, JP Hoffmann, RG Wood, JD Hudetz, AG Hillery, CA
Liao JK, Zulueta JJ, Yu FS, Peng HB, Cote CG, Hassoun PM: Regulation of bovine endothelial constitutive nitric oxide synthase by oxygen. J Clin Invest 1995; 96:2661–6Liao, JK Zulueta, JJ Yu, FS Peng, HB Cote, CG Hassoun, PM
Phelan MW, Faller DV: Hypoxia decreases constitutive nitric oxide synthase transcript and protein in cultured endothelial cells. J Cell Physiol 1996; 167:469–76Phelan, MW Faller, DV
Reiter CD, Wang X, Tanus-Santos JE, Hogg N, Cannon RO III, Schechter AN, Gladwin MT: Cell-free hemoglobin limits nitric oxide bioavailability in sickle-cell disease. Nat Med 2002; 8:1383–9Reiter, CD Wang, X Tanus-Santos, JE Hogg, N Cannon, RO Schechter, AN Gladwin, MT
Gladwin MT, Schechter AN, Ognibene FP, Coles WA, Reiter CD, Schenke WH, Csako G, Waclawiw MA, Panza JA, Cannon RO III: Divergent nitric oxide bioavailability in men and women with sickle cell disease. Circulation 2003; 107:271–8Gladwin, MT Schechter, AN Ognibene, FP Coles, WA Reiter, CD Schenke, WH Csako, G Waclawiw, MA Panza, JA Cannon, RO
Aslan M, Freeman BA: Oxidant-mediated impairment of nitric oxide signaling in sickle cell disease: Mechanisms and consequences. Cell Mol Biol (Noisy-le-grand) 2004; 50:95–105Aslan, M Freeman, BA
Martinez-Ruiz R, Montero-Huerta P, Hromi J, Head CA: Inhaled nitric oxide improves survival rates during hypoxia in a sickle cell (SAD) mouse model. Anesthesiology 2001; 94:1113–8Martinez-Ruiz, R Montero-Huerta, P Hromi, J Head, CA
Head CA, Brugnara C, Martinez-Ruiz R, Kacmarek RM, Bridges KR, Kuter D, Bloch KD, Zapol WM: Low concentrations of nitric oxide increase oxygen affinity of sickle erythrocytes in vitro  and in vivo  . J Clin Invest 1997; 100:1193–8Head, CA Brugnara, C Martinez-Ruiz, R Kacmarek, RM Bridges, KR Kuter, D Bloch, KD Zapol, WM
Gladwin MT, Schechter AN, Shelhamer JH, Pannell LK, Conway DA, Hrinczenko BW, Nichols JS, Pease-Fye ME, Noguchi CT, Rodgers GP, Ognibene FP: Inhaled nitric oxide augments nitric oxide transport on sickle cell hemoglobin without affecting oxygen affinity. J Clin Invest 1999; 104:937–45Gladwin, MT Schechter, AN Shelhamer, JH Pannell, LK Conway, DA Hrinczenko, BW Nichols, JS Pease-Fye, ME Noguchi, CT Rodgers, GP Ognibene, FP
Lopez BL, Barnett J, Ballas SK, Christopher TA, Davis-Moon L, Ma X: Nitric oxide metabolite levels in acute vaso-occlusive sickle-cell crisis. Acad Emerg Med 1996; 3:1098–103Lopez, BL Barnett, J Ballas, SK Christopher, TA Davis-Moon, L Ma, X
Sullivan KJ, Kissoon N, Duckworth LJ, Sandler E, Freeman B, Bayne E, Sylvester JE, Lima JJ: Low exhaled nitric oxide and a polymorphism in the NOS I gene is associated with acute chest syndrome. Am J Respir Crit Care Med 2001; 164:2186–90Sullivan, KJ Kissoon, N Duckworth, LJ Sandler, E Freeman, B Bayne, E Sylvester, JE Lima, JJ
Powars DR, Conti PS, Wong WY, Groncy P, Hyman C, Smith E, Ewing N, Keenan RN, Zee CS, Harold Y, Hiti AL, Teng EL, Chan LS: Cerebral vasculopathy in sickle cell anemia: Diagnostic contribution of positron emission tomography. Blood 1999; 93:71–9Powars, DR Conti, PS Wong, WY Groncy, P Hyman, C Smith, E Ewing, N Keenan, RN Zee, CS Harold, Y Hiti, AL Teng, EL Chan, LS
Nuutinen J, Liu Y, Laakso MP, Karonen JO, Roivainen R, Vanninen RL, Partanen K, Ostergaard L, Sivenius J, Aronen HJ: Assessing the outcome of stroke: A comparison between MRI and clinical stroke scales. Acta Neurol Scand 2006; 113:100–7Nuutinen, J Liu, Y Laakso, MP Karonen, JO Roivainen, R Vanninen, RL Partanen, K Ostergaard, L Sivenius, J Aronen, HJ
Powars DR: Management of cerebral vasculopathy in children with sickle cell anaemia. Br J Haematol 2000; 108:666–78Powars, DR
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx). 
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx). 
Fig. 1. Plasma nitric oxide metabolite levels before inhaled nitric oxide (INO) and at 3 and 22 h after therapy. The  dashed line  represents the normal/control level of plasma nitric oxide metabolites (NOx).