Erythrocyte type 1 equilibrative nucleoside transporter expression in sickle cell disease and sickle cell trait.

Hypoxia-mediated red blood cell (RBC) sickling is central to the pathophysiology of sickle cell disease (SCD). The signalling nucleoside adenosine is thought to play a significant role in this process. This study investigated expression of the erythrocyte type 1 equilibrative nucleoside transporter (ENT1), a key regulator of plasma adenosine, in adult patients with SCD and carriers of sickle cell trait (SCT). Relative quantitative expression analysis of erythrocyte ENT1 was carried out by Western blot and flow cytometry. Patients with SCD with steady state conditions, either with SS or SC genotype, untreated or under hydroxycarbamide (HC) treatment, exhibited a relatively high variability of erythrocyte ENT1, but with levels not significantly different from normal controls. Most strikingly, expression of erythrocyte ENT1 was found to be significantly decreased in patients with SCD undergoing painful vaso-occlusive episode and, unexpectedly, also in healthy SCT carriers. Promoting hypoxia-induced adenosine signalling, the reduced expression of erythrocyte ENT1 might contribute to the pathophysiology of SCD and to the susceptibility of SCT individuals to altitude hypoxia or exercise to exhaustion.

Lipid profiles in French West Indies sickle cell disease cohorts, and their general population.

BACKGROUND: The pathophysiology of sickle cell disease (SCD) and the variability of its clinical expression remain not fully understood, whether within or between different SCD genotypes. Recent studies have reported associations between lipid levels and several SCD complications. If lipid levels have been previously described as low in sickle cell anemia (SCA), few data have been provided for sickle cell SC disease (SCC). We designed our epidemiological study to isolate lipid levels and profiles by genotype in Guadeloupian cohorts of SCA and SCC adult patients, at steady state. We compared SCD lipid levels with those of the Guadeloupian general population (GGP), and analyzed potential associations between lipid levels and SCD complications (vaso-occlusive crises, acute chest syndrome and osteonecrosis). METHODS: Lipids, apolipoproteins, biological variables and anthropometric evaluation, were collected at steady state from medical files for 62 SCC and 97 SCA adult patients. Clinical SCD complications were collected from the clinical files. Analysis was conducted by genotype for all variables. RESULTS: Different SCC and SCA lipid profiles, both distinct from their GGP's, were identified. Compared to SCC and GGP, higher triglyceride (TG) levels were observed in SCA patients, independent of hydroxyurea, hemolysis, gender, age, body mass index (BMI), abdominal obesity and clinical nutritional status. Our survey highlights also subsequent anthropometrical phenotypes, with an over-representation of abdominal obesity with normal BMI in SCA patients, and affecting almost exclusively females in both genotypes. Moreover, more frequent positive history of acute chest syndrome (ACS) was observed in SCA patients with TG level higher than 1.50 g/l, and of osteonecrosis in SCC patients having non high-density lipoprotein-cholesterol level (Non HDL-C) higher than 1.30 g/l. CONCLUSIONS: This study reveals that SCA and SCC patients exhibit distinct lipid profiles and suggests that high TG and Non HDL-C levels are associated with past histories of ACS and osteonecrosis in SCA and SCC patients, respectively.

Association between oxidative stress and vascular reactivity in children with sickle cell anaemia and sickle haemoglobin C disease.

Oxidative stress and haemolysis-associated nitric oxide (NO) depletion plays a crucial role in the development of vasculopathy in sickle cell anaemia (SS). However it remains unknown whether oxidative stress and haemolysis levels influence vascular function in patients with sickle haemoglobin C disease (SC). Microvascular response to heat (using Laser Doppler flowmetry on finger), oxidative stress biomarkers, NO metabolites, endothelin-1 and haematological parameters were compared between patients with SS and SC. Vascular function, oxidative and nitrosative markers were also measured in healthy (AA) children. SS and SC had increased plasma advanced oxidation protein products (AOPP), malondialdehyde, plasma antioxidant activities and NO end products, compared to AA. SC had lower catalase activity compared to AA and SS. Haemolytic rate, glutathione peroxidase and nitrotyrosine concentrations were significantly increased in children with SS compared to SC and AA. SS and SC had impaired microvascular reactivity compared to AA. In SS, the plateau phase of the response to local thermal heating was negatively associated with nitrotyrosine and AOPP. No association between vascular function parameters and oxidative stress markers was observed in SC. Mild haemolysis in SC, compared to SS, may limit oxidative and nitrosative stress and could explain the better preserved microvascular function in this group.

Exacerbation of oxidative stress during sickle vaso-occlusive crisis is associated with decreased anti-band 3 autoantibodies rate and increased red blood cell-derived microparticle level: a prospective study.

Painful vaso-occlusive crisis, a hallmark of sickle cell anaemia, results from complex, incompletely understood mechanisms. Red blood cell (RBC) damage caused by continuous endogenous and exogenous oxidative stress may precipitate the occurrence of vaso-occlusive crises. In order to gain insight into the relevance of oxidative stress in vaso-occlusive crisis occurrence, we prospectively compared the expression levels of various oxidative markers in 32 adults with sickle cell anaemia during vaso-occlusive crisis and steady-state conditions. Compared to steady-state condition, plasma levels of free haem, advanced oxidation protein products and myeloperoxidase, RBC caspase-3 activity, as well as the concentrations of total, neutrophil- and RBC-derived microparticles were increased during vaso-occlusive crises, whereas the reduced glutathione content was decreased in RBCs. In addition, natural anti-band 3 autoantibodies levels decreased during crisis and were negatively correlated with the rise in plasma advanced oxidation protein products and RBC caspase-3 activity. These data showed an exacerbation of the oxidative stress during vaso-occlusive crises in sickle cell anaemia patients and strongly suggest that the higher concentration of harmful circulating RBC-derived microparticles and the reduced anti-band 3 autoantibodies levels may be both related to the recruitment of oxidized band 3 into membrane aggregates.

Micro- and macrovascular function in children with sickle cell anaemia and sickle cell haemoglobin C disease.

It is unclear whether vascular function is affected similarly in children with sickle cell anaemia (SS) and children with sickle haemoglobin C (SC) disease. Therefore, we compared micro and macrovascular functions in healthy (AA) children, children with SS and SC disease, and assessed their association with physical activity. Participants (24 SS, 22 SC and 16 AA), were compared in terms of 1) thermal hyperaemic response (finger pad warming to 42°C) measured by Laser Doppler techniques, 2) arterial stiffness determined by pulse wave velocity, 3) daily energy expenditure related to moderate and intense physical activities estimated by questionnaire and 4) fitness level, evaluated by the six-minute walk test. Response to heating differed between SS, SC and controls. Peripheral microvascular reactivity was lower and pulse wave velocity higher in SS compared to AA. SC had blunted microvascular reactivity in response to heating compared to AA but pulse wave velocity was not different within the two groups. Physical activity and fitness levels were markedly lower in sickle cell patients compared to healthy controls but no association was observed with vascular function. Microvasodilatory reserve is decreased in both SS and SC patients but only SS patients were also characterised by impaired macrovascular function.

Cerebral and muscle microvascular oxygenation in children with sickle cell disease: Influence of hematology, hemorheology and vasomotion.

The present study investigated cerebral and muscle hemoglobin oxygen saturation (tissue oxygen index, TOI) in children with sickle cell anemia (SS), sickle cell hemoglobin C disease (SC) and healthy children (AA). TOI was measured by near-infrared spectroscopy (NIRS) and spectral analysis of the TOI variability was used to assess flowmotion and vasomotion. Arterial oxyhemoglobin saturation (SpO2), hemorheological and hematological parameters were also measured in SS and SC children. Both TOI were lower in SS compared to both AA and SC children, with SC exhibiting lower values than AA children. Cerebral vasomotion expressed in absolute values was enhanced in SS compared to AA and SC children. Muscle vasomotion did not differ between the three groups. Hematocrit, SpO2 and red blood cell deformability were positively associated with cerebral TOI in SS children. We demonstrated that 1) cerebral and muscle TOI were markedly decreased in SS children while the decrease of TOI was milder in SC children, 2) cerebral TOI level was associated with several biological markers in SS children only and 3) cerebral vasomotion was enhanced in SS, possibly to counterbalance the effects of chronic cerebral hypoxia.

Association of adenylyl cyclase 6 rs3730070 polymorphism and hemolytic level in patients with sickle cell anemia.

A recent study suggested that adenosine signaling pathway could promote hemolysis in patients with sickle cell anemia (SCA). This signaling pathway involves several gene coding enzymes for which variants have been described. In this study, we analyzed the genotype-phenotype relationships between functional polymorphisms or polymorphisms associated with altered expression of adenosine pathway genes, namely adenosine deaminase (ada; rs73598374), adenosine A2b receptor (adora2b; rs7208480), adenylyl cyclase6 (adcy6; rs3730071, rs3730070, rs7300155), and hemolytic rate in SCA patients. One hundred and fifty SCA patients were genotyped for adcy6, ada, and adora2b variants as well as alpha-globin gene, a genetic factor known to modulate hemolytic rate. Hematological and biochemical data were obtained at steady-state. Lactate dehydrogenase, aspartate aminotransferase, reticulocytes and total bilirubin were used to calculate a hemolytic index. Genotype-phenotype relationships were investigated using parametric tests and multivariate analysis. SCA patients carrying at least one allele of adcy6 rs3730070-G exhibited lower hemolytic rate than non-carriers in univariate analysis (p=0.006). The presence of adcy6 rs3730070-G variant was associated with a decreased hemolytic rate in adjusted model for age and alpha-thalassemia (p=0.032). Our results support a protective effect of adcy6 rs3730070-G variant on hemolysis in SCA patients.

Which side of the balance determines the frequency of vaso-occlusive crises in children with sickle cell anemia: Blood viscosity or microvascular dysfunction?

Vascular resistance and tissue perfusion may be both affected by impaired vascular function and increased blood viscosity. Little is known about the effects of vascular function on the occurrence of painful vaso-occlusive crises (VOC) in children with sickle cell anemia (SCA). The aim of the present study was to determine which side of the balance (blood viscosity or vascular function) is the most deleterious in SCA and increases the risk for frequent hospitalized VOC. Microvascular function, microcirculatory oxygenation and blood viscosity were determined in a group of 22 SCA children/adolescents at steady state and a group of 13 healthy children/adolescents. Univariate analyses demonstrated blunted microvascular reactivity during local thermal heating test and decreased microcirculatory oxygenation in SCA children compared to controls. Multivariate analysis revealed that increased blood viscosity and decreased microcirculatory oxygenation were independent risk factors of frequent VOC in SCA. In contrast, the level of microvascular dysfunction does not predict VOC rate. In conclusion, increased blood viscosity is usually well supported in healthy individuals where vascular function is not impaired. However, in the context of SCA, microvascular function is impaired and any increase of blood viscosity or decrease in microcirculatory oxygenation would increase the risks for frequent VOC.

Physical activity level is not a determinant of autonomic nervous system activity and clinical severity in children/adolescents with sickle cell anemia: A pilot study.

BACKGROUND: Autonomic nervous system (ANS) activity has been suggested to modulate the clinical severity of sickle cell anemia (SCA) by increasing the risk for vaso-occlusive events. Regular physical activity (PA) is known to improve ANS activity and health status in several cardiovascular and metabolic diseases. Whether regular PA improves the health status of SCA patients remains unknown. PROCEDURE: Twenty-two patients with SCA and 15 healthy (AA) children/adolescents participated to the study. Heart rate variability was measured in supine position and after a tilt-test to quantify the ANS activity. PA energy expenditure (PAEE) was assessed with questionnaire. RESULTS: 1) PAEE was lower in SCA compared to AA (190 ± 152 vs. 432 ± 277 kcal · d(-1), respectively, P < 0.01), 2) overall ANS activity was lower in SCA compared to AA, 3) parasympathetic withdrawal was observed in SCA with aging, 4) ANS reactivity was slightly impaired in SCA compared to AA (reduction in HFnu: -38 ± 27 vs. -58 ± 14%, respectively, P < 0.05), 5) ANS indices, PAEE, and rates of clinical events were not correlated. CONCLUSION: Both the level of PA and ANS activity are reduced in SCA compared to AA children/adolescents, particularly in those older than 15 years. Neither PAEE, nor ANS activity seem to influence the clinical severity of children/adolescents with SCA.

Hematologic and hemorheological determinants of resting and exercise-induced hemoglobin oxygen desaturation in children with sickle cell disease.

The aim of the study was to determine the factors associated with resting and exercise-induced hemoglobin oxygen desaturation. The well-established six-minute walk test was conducted in 107 sickle cell children (50 with sickle hemoglobin C disease and 57 with sickle cell anemia) at steady state. Hemoglobin oxygen saturation was measured before and immediately after the six-minute walk test. Blood samples were obtained on the same day to measure hematologic and hemorheological parameters. Exercise-induced hemoglobin oxygen desaturation was defined as a drop in hemoglobin oxygen saturation of 3% or more at the end of the six-minute walk test compared to resting levels. No children with sickle hemoglobin C disease, but approximately 50% of children with sickle cell anemia showed mild or moderate oxygen desaturation at rest, which was independently associated with the percentage of reticulocytes. Exercise-induced hemoglobin oxygen desaturation was observed in 18% of children with sickle hemoglobin C disease and 34% of children with sickle cell anemia, and was independently associated with the six-minute walk test, acute chest syndrome rate and the strength of red blood cell aggregates in children with sickle cell anemia. No association was found in children with sickle hemoglobin C disease between exercise-induced hemoglobin oxygen desaturation and the measured parameters. Hemoglobin oxygen desaturation at rest was common in children with sickle cell anemia but not in children with sickle hemoglobin C disease, and was mainly associated with greater hemolysis. Physiological strain during exercise and red blood cell aggregation properties may predict the occurrence of exercise-induced hemoglobin oxygen desaturation in children with sickle cell anemia.