Melville Q. Wyche

Nitric oxide and cyclic GMP levels in sickle cell patients receiving hydroxyurea

Summary. Recent studies suggest that nitric oxide (NO) may partly be responsible for the beneficial effect of hydroxyurea (HU) in sickle cell disease (SCD) patients. NO stimulates cyclic guanosine monophosphate (cGMP) production, which mediates vasodilatation. We investigated the association between NO, cGMP and fetal haemoglobin (HbF) levels after HU administration. Our data showed that chronic HU significantly increased NO, cGMP, and HbF levels in SCD. Recently it was shown that HbF production was stimulated by cGMP‐dependent protein kinase. Our results suggest that NO stimulates cGMP production, which then activates a protein kinase and increases the production of HbF.

Nitric Oxide Metabolites in Sickle Cell Anemia Patients after Oral Administration of Hydroxyurea

The mechanism of action of hydroxyurea (HU) in decreasing the frequency of pain crisis in sickle cell disease (SCD) has not been fully elucidated. In vitro and in vivo studies suggest that nitric oxide (NO), a potent vasodilator, may partly be responsible for the beneficial effect of HU. This study was designed to determine the effect of oral administration of HU on plasma levels of NO metabolites (NOx) in sickle cell patients (SCP). The results indicate that during steady-state plasma levels of NOx were significantly higher in HU-treated patients compared to non HU-treated patients or normal controls (p <.05). In five inpatients in mild pain plasma levels of NOx increased significantly after 2h of HU administration (p <.05); however, in three inpatients in persistent pain with significantly lower baseline NOx there was a minimal NOx response to HU at 2h (p <.01). These observations indicate that HU administration is associated with the production of NO in some SCP, but that further study of the pharmacodynamics of this effect is necessary.

Cerebral oximetry in patients with sickle cell disease

Background  There is limited information concerning the brains oxygen supply and demand in patients with sickle cell disease.

Effects of hydroxyurea and L-arginine on the production of nitric oxide metabolites in cultures of normal and sickle erythrocytes

Abstract Previous in vitro studies suggest that erythrocytes may be a source of nitric oxide (NO) produced by nitric oxide synthase (NOS) or by oxyhemoglobin-mediated oxidation of hydroxyurea (HU). This study was performed to determine the roles of HU and NOS in the production of NO by normal and sickle erythrocytes. Red blood cells (RBCs) from normal adult hemoglobin (HbAA) and homozygous sickle cell subjects (HbSS) were incubated with PBS containing 0.2 mM hydrogen peroxide (control) for 2 h at 37°C in the presence and absence of L-arginine, the substrate for NOS, and with L-arginine plus HU in the presence and absence of L-NMMA, a specific inhibitor of NOS. The nitrate and nitrite metabolites of NO, expressed as [NOx], were measured. [NOx] in the HbAA and HbSS RBC cultures was not significantly different in the presence and absence of 1.0 mM L-arginine (p > 0.1). [NOx] in the HbAA and HbSS cultures treated with a clinically relevant dose of HU (1.0 mM) plus 1.0 mM L-arginine was significantly greater than that in controls incubated with PBS and with L-arginine p < 0.01. However, [NOx] in the HbAA and HbSS cultures treated with 50 μg/ml L-NMMA was not significantly different than that in the cultures treated with HU plus L-arginine in the absence of L-NMMA. These findings suggest that NOx production by erythrocytes may be increased by treatment with HU and may not be decreased by inhibiting NOS. Therefore, we conclude that a therapeutic dose of HU may increase the plasma concentration of NO by a mechanism that does not require erythrocytes NOS activity

Plasma Levels of TNF- in Sickle Cell Patients Receiving Hydroxyurea

Abstract Hydroxyurea (HU), a chemotherapeutic agent, used increasingly in the treatment of sickle cell disease (SCD) stimulates the release of a tumor necrosis factor (TNF-) from human macrophages in vitro and the concentration of TNF- is greater than normal in subjects affected by SCD. It is widely accepted that HU may inhibit vaso-occlusive crisis (VOC) by stimulating the production of fetal hemoglobin (HbF) and nitric oxide (NO) in SCD; however, the beneficial effects of HU in vivo may be counteracted by the release of TNF- and, in turn, the expression of a vascular cell adhesion molecule (VCAM-1) on leukocytes. Previous studies have shown that the severity of SCD increases with the leukocyte count. Therefore, we examined the relationship between plasma levels of TNF- and HbF in SCD patients during steady-state (StSt) conditions (in the absence of VOC) and during VOC conditions after the acute administration of HU. Venous blood was collected in SCD patients over 6 h after administering a single dose of HU. Plasma TNF- was found to be greater in SCD subjects than in reported normal adult controls (p<0.05). TNF- in the StSt group was not significantly different than in the VOC group; however, the plasma TNF- tended to greater in the VOC group (p>0.1). An increase in the HbF concentration after acute administration of HU (p<0.01) was not associated with a significant change in plasma TNF- (p>0.1). Contrary to the results of in vitro studies, HU did not increase the plasma concentration of TNF-. These findings suggest that a HU-induced increase in TNF- does not contribute to VOC and sickle cell patients can be counseled that the HU-induced increase in TNF- does not counteract the beneficial effects of HU in SCD.

Effect of transfusion on cerebral oxygenation, flow velocity in a patient with sickle cell anemia and Moyamoya disease: a case report.

Abstract Vascular occlusive diseases affect brain blood flow, brain metabolism and are associated with arterial ischemic stroke. This study was designed to measure the brain blood flow velocity, brain oxygenation, hemoglobin concentrations, hematocrit, and cell free hemoglobin at pre- and post-exchange red cell transfusion in an 18 year old male patient with sickle cell disease and moyamoya syndrome (MMS). Exchange transfusion increased cerebral oxygen saturation 12%, total hemoglobin concentration 2%, hemoglobin AA 80%, and reduced sickle (SS) hemoglobin 12%, arterializations 33%, and cell free hemoglobin 33%. Brain blood flow velocity values were unaffected by transfusion. These observations suggest that exchange transfusion increases the hemoglobin carrying capacity and reduces sickle hemoglobin and shunting of blood, which may improve the peripheral and cerebral oxygenation. Transfusion did not affect the brain blood flow in this patient. Therefore the risk of transient ischemic attack and arterial ischemic stroke from mms still exist.

Arterialization of Venous Blood for Differentiation of Sickle Cell Subjects in Vaso-occlusive Crisis

Abstract These studies were designed as two experiments. Experiment 1 was performed to validate the hypothesis that oxygen saturation of the venous blood may be a marker for vaso-occlusive crisis (VOC) in sickle cell patients undergoing hydroxyurea (HU) treatments. Experiment 2 was performed to test the hypothesis that an acute increase in the blood nitric oxide (NO) concentration by administering HU modulates the perception of pain in sickle cell subjects in VOC. The percent saturations of oxyhemoglobin (%O2Hb), reduced hemoblogin (%RHb), carboxy-hemoglobin (%COHb), met-hemoglobin (%MHb), fetal hemoglobin (HbF), and nitric oxide metabolites were measured in venous blood samples collected from sickle cell disease (SCD) who were on and off HU and O2 at steady state and during VOC. The results showed the ratio of %O2Hb/RHb in VOC+HU was significantly higher than patients in the steady state who were on and off of HU (p<0.05). The %COHb was higher in all SCD groups, %COHb values were significantly different in SCD at steady state who were on HU. HU and O2 treatment did not play important role on venous blood %O2Hb and pain scores in SCD during VOC. A single oral dose of HU was associated with a significant increase in the venous concentration of nitric oxide metabolites (NOx), p<0.05. These findings suggest that the ratio %O2Hb/RHb in venous blood and pain scores differentiate HU-untreated and HU-treated at steady state subjects from HU-treated subjects in VOC; however, the acute increase in venous NOx produced by administering HU to HU-treated subjects in VOC does not explain this difference.