Fetal Hemoglobin Concentration Research Articles

Osmotic parameters of red blood cells from umbilical cord blood

The transfusion of red blood cells from umbilical cord blood (cord RBCs) is gathering significant interest for the treatment of fetal and neonatal anemia, due to its high content of fetal hemoglobin as well as numerous other potential benefits to fetuses and neonates. However, in order to establish a stable supply of cord RBCs for clinical use, a cryopreservation method must be developed. This, in turn, requires knowledge of the osmotic parameters of cord RBCs. Thus, the objective of this study was to characterize the osmotic parameters of cord RBCs: osmotically inactive fraction (b), hydraulic conductivity (Lp), permeability to cryoprotectant glycerol (Pglycerol), and corresponding Arrhenius activation energies (Ea). For Lp and Pglycerol determination, RBCs were analyzed using a stopped-flow system to monitor osmotically-induced RBC volume changes via intrinsic RBC hemoglobin fluorescence. Lp and Pglycerol were characterized at 4°C, 20°C, and 35°C using Jacobs and Stewart equations with the Ea calculated from the Arrhenius plot. Results indicate that cord RBCs have a larger osmotically inactive fraction compared to adult RBCs. Hydraulic conductivity and osmotic permeability to glycerol of cord RBCs differed compared to those of adult RBCs with the differences dependent on experimental conditions, such as temperature and osmolality. Compared to adult RBCs, cord RBCs had a higher Ea for Lp and a lower Ea for Pglycerol. This information regarding osmotic parameters will be used in future work to develop a protocol for cryopreserving cord RBCs.

Methemoglobin measure in adult patients with sickle-cell anemia: influence of hydroxyurea therapy

Introduction: Hemoglobin S (HbS) is unstable hemoglobin that easily oxidizes, causing methemoglobin (MetHb) increased production in patients with sickle-cell anemia (SCA). Objectives: To determine MetHb levels and the influence of hydroxyurea (HU) therapy on this marker in patients with SCA. Materials and methods: Blood samples from 53 patients with SCA at the steady-state, with and without HU therapy, and 30 healthy individuals were collected to evaluate MetHb levels. The MetHb measurement was performed by spectrophotometry. Complete blood count, HU measurements, and fetal hemoglobin (HbF) and HbS concentrations were taken from medical records. Results: MetHb levels were statically higher in patients with SCA when compared to control group (p < 0.001). There was no statistical difference in MetHb level between SCA patients, either using or not HU. We obtained a positive correlation between MetHb measurements and HbS concentration (r = 0.2557; p = 0.0323). Conclusion: HbS presence favored hemoglobin breaking down, and consequently increased MetHb production. Treatment with HU, however, did not influence the levels of this marker.

Intrauterine Transfusions in FHD: When and How?

ABSTRACT Intrauterine fetal blood transfusions still remain the gold standard of prenatal therapy in severe cases of fetal hemolytic disease due to mother-fetus immunization. Middle cerebral artery-peak systolic velocity (MCA-PSV) measurements plays the most important role in diagnosing the disease. A value of MCA-PSV &gt;1.5 allows us to diagnose severe or moderate anemia and prompts us to treat the patient. The time of subsequent transfusions is estimated by the hemoglobin level directly after the transfusion and the fact that the concentration of fetal hemoglobin in blood decreases at a rate of 0.3 g% per day. Even though effective, these procedures carry with them major risks and that is why prophylaxis is essential. How to cite this article Wielgos M, Brawura-Biskupski- Samaha R. Intrauterine Transfusions in FHD: When and How? Donald School J Ultrasound Obstet Gynecol 2013;7(4):481-483.

The Effects Of The Histone Methyltransferase Inhibitor UNC0638 Upon Gamma Globin Gene and Protein Expression Are Erythroblast Differentiation Stage Specific

Epigenetic modification of chromatin in erythroid cells represents an active field of study aimed, in part, toward increased expression of fetal hemoglobin in patients with beta-thalassemia. The homologous methyltransferases G9a and GLP regulate globin gene transcription by catalyzing mono- and dimethylation at Lys 9 and dimethylation at Lys 27 of histone H3. Inhibition of these methyltransferases by the small molecule named UNC0638 was recently shown to increase gamma-globin gene expression in adult human hematopoietic precursor and stem cells. Here UNC0638 was explored further to include fetal hemoglobin expression among more mature erythroid cells cultured from CD34(+) cells of three healthy adult human donors in a serum-free culture medium. According to this culture model, the main erythroblast population on culture days 0-7 consists of CD36(+), CD45(+), CD71(moderate), CD235a(-) erythroid progenitor cell. On culture days 7-14, the progenitor cells differentiate in the presence of erythropoietin to become CD36(+), CD45(-), CD71(high), CD235a(+) precursor cells. During the final week in culture, the erythroblasts undergo nuclear condensation, enucleation, and loss of RNA combined with the loss of CD36 and CD71 on the plasma membrane to become mature erythrocytes. To investigate different stages of erythroblast maturation, the cells were cultured in medium containing 1µM UNC0638 for periods of seven days (culture days 0-7, 7-14, or 14-21) and compared to control cultures without UNC0638. The effects of UNC0638 were determined by flow cytometry, Q-RT-PCR and hemoglobin chromatography (HPLC).Unexpectedly, fetal hemoglobin expression was highly-dependent upon the differentiation stage of the cells in the presence of UNC0638. When cultured in UNC0638 supplemented medium on culture days 0-7 or 14-21, the cells underwent terminal maturation, but there was no significant increase in the fetal hemoglobin content of the mature cells (see abstract figure). In contrast, UNC0638 added on culture days 7-14, caused a significant increase in fetal hemoglobin (HbF; control: 3.9 ± 3.5% vs. day 7-14 UNC0638: 32.6 ± 0.95%, p=0.007). The increase in HbF was associated with a similar increase in gamma-globin mRNA (control: 1.5E+06 ± 1.7E+05 copies/ng vs. day 7-14 UNC0638: 7.5E+06 ± 1.4E+06 copies/ng, p=0.021). Additionally, terminal maturation and enucleation were partially inhibited when compared to the other conditions or controls.These data suggest that UNC0638 causes a robust increase in fetal hemoglobin as the cells undergo maturation. Fetal hemoglobin increases were more pronounced after exposure to UNC0638 during the erythropoietin-dependent transition from CD235a(-) to CD235a(+) erythroblasts. The results suggest that fetal hemoglobin regulation by G9a and GLP may be differentiation stage dependent. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

The use of cell salvage in women undergoing cesarean hysterectomy for abnormal placentation

BackgroundAlthough transfusion of autologous blood obtained from cell salvage has increased, its role in obstetric practice remains controversial. This case series reports the use of cell salvage in an attempt to avoid allogeneic transfusion in women undergoing cesarean hysterectomy for placenta accreta. MethodsThis prospective observational study, conducted in a large public maternity hospital, included 41 women with an antenatal diagnosis of placenta accreta, of whom 20 underwent cesarean hysterectomy and 15 received autologous blood after cell salvage. Intraoperative cell salvage was used for autologous blood transfusion, and salvaged blood was monitored for prewash and postfiltration squamous cells, fetal hemoglobin, and potassium concentration. Pre- and postoperative hemoglobin, platelet count and coagulation profile were compared. ResultsTwenty women underwent caesarean hysterectomy. Cell-salvaged blood was collected in 18 women and re infused in 15 women (83.3%). The mean volume of reinfused salvaged blood was 1476±247mL. Mean potassium concentrations (1.4±1.2 versus 3.7±0.42mEq/L) and median squamous cell counts (0 [0–1] versus 8 [3–12]/high power field) were significantly lower postfiltration compared to prewash values. There were no instances of intraoperative or postoperative amniotic fluid embolism, hypotension, sepsis or coagulopathy. Of the 15 women who received autologous blood, 13 (86.7%) did not require allogeneic red blood cell transfusion. ConclusionsAutologous transfusion of salvaged blood can be used to minimize allogeneic transfusion in women undergoing cesarean hysterectomy for placenta accreta.

PF.25 Relationship Between Maternal Antibody Level and Fetal Haemoglobin Concentration in Pregnancies Complicated by Rhesus-D Alloimmunization

BackgroundCurrently, maternal Rh-D antibody levels are primarily used to triage which alloimmunized women warrant enhanced surveillance with middle cerebral artery Doppler. Traditionally, maternal Rh-D antibody levels ≤15 IU/ml have indicated,...

Fetal hemoglobin and hydroxycarbamide moduate both plasma concentration and cellular origin of circulating microparticles in sickle cell anemia children

Microparticles are cell membrane-derived microvesicles released during cell apoptosis and activation processes. They have been described as bio-markers in various vascular diseases, including sickle cell anemia, and associated with an increased risk of thrombosis. We investigated the effects of fetal hemoglobin level, a factor known to modulate the clinical expression of sickle cell anemia, and that of hydroxycarbamide treatment which reduces the frequency of vasoocclusive crises, the canonical clinical manifestation of the disease, on both the plasma concentration and the cellular origin of circulating microparticles. Flow cytometry was used to characterize microparticles in 62 sickle cell anemia children at steady state aged 2 months-16 years; 13 of them were treated with hydroxycarbamide. In untreated children, we observed negative correlations between fetal hemoglobin levels and the absolute plasma concentration of microparticles as well as that of microparticles specifically derived from platelets, erythrocytes, and monocytes. Compared to untreated children, those treated with hydroxyurea showed lower concentrations of total microparticles as a consequence of decreased microparticles shed by platelets and erythrocytes. In conclusion, in our sickle cell patients, neonatal decline of fetal hemoglobin coincided with an increase in circulating microparticles derived from erythrocytes, platelets, and monocytes. Hydroxyurea treatment was associated with a decrease in microparticles derived from erythrocytes and platelets.

561: Relationship between maternal antibody level and fetal hemoglobin concentration in pregnancies complicated by Rh-D alloimmunization

Background Currently, maternal Rh-D antibody levels are primarily used to triage which alloimmunized women warrant enhanced surveillance with middle cerebral artery Doppler. Traditionally, maternal Rh-D antibody levels ≤15 IU/ml have indicated, at worst, mild anaemia and provided reassurance. This threshold has not been widely studied. Methods A prospective cohort study of all intrauterine fetal transfusions (IUT) for Rh-D alloimmunization performed at our tertiary fetal medicine unit from 1996–2011. Fetal haemoglobin (Hb) levels at the time of IUT were adjusted for gestational age (multiples of median [MoM]) and correlated with the maternal serum Rh-D antibody level taken on the day of IUT, or ≤2 weeks prior to the transfusion. Results 260 IUTs were performed, of which 195 were for Rh-D alloimmunzation in 82 pregnancies. No significant correlation was demonstrated between fetal Hb and serum antibody levels (Spearman r = 0.08; p = 0.35). Rates of mild (0.65–0.84 MoM), moderate (0.55–0.64 MoM) and severe ( 15 IU/ml for detecting any fetal anaemia ( 15 IU/ml detecting moderate-severe anaemia ( 8 IU/ml, the sensitivity, specificity, PPV and NPV of maternal serum antibody levels detecting any fetal anaemia were 96%, 5%, 85% and 17% respectively. Conclusion The widely used Rh-D threshold of >15 IU/ml may miss a substantial proportion of cases of fetal anaemia.

Editorial Commentary: Intermittent Preventive Treatment in Pregnancy With Sulfadoxine-Pyrimethamine: The Controversy Continues

Editorial Commentary: Intermittent Preventive Treatment in Pregnancy With Sulfadoxine-Pyrimethamine: The Controversy Continues

OS089. Elevated levels of the heme scavenger alpha-1-microglobulin in maternal plasma at the end of first trimester in patients who subsequently develop preeclampsia

Resent research has revealed an increased concentration of free fetal hemoglobin (HbF) in maternal serum from patients who subsequently develops preeclampsia (PE). In a previous study of 96 patients we have shown that HbF in combination with the heme-scavenger alpha-1-microglobulin (A1M) are potential predictive biomarkers of PE. In this validating case-control study we aimed to confirm the previous findings, that A1M is elevated in the maternal circulation at the end of first trimester in patients who subsequently develops PE. In this study A1M was measured in plasma instead of serum. Patients were recruited from an ongoing prospective study of new biomarkers to predict and diagnose PE. In total we included 84 patients. 8 patients subsequently developed PE, 4 developed pregnancy induced hypertension (PIH) and 72 were controls with uncomplicated pregnancies. The plasma samples were all taken at 7+0-18+0 weeks of gestation (mean 12+1) and analyzed for concentrations of A1M with Radioimmuno Assay (RIA). This method has been previously described in details. Statistics was performed using one-way ANOVA. The mean plasma concentration of A1M in the PE group was 8.6mg/ml, 6.0 in the PIH group and 7.1mg/ml in the controls group. The PE group differed significantly from the controls group (p=0.004), whereas the PIH group did not differ significantly from the controls. Our findings in plasma confirm previous findings described for serum, i.e. A1M is significantly increased in in first trimester maternal plasma in patients who subsequently develops PE. Since A1M is the most efficient heme scavenger we suggest that A1M may be a physiological defense mechanism against the elevated levels of free HbF found in patients who subsequently develops PE or in patients with manifest PE. Furthermore, A1M did not increase in patients who develops PIH later in their pregnancies indicating its specificity for PE.

Genetic modifiers of sickle cell disease

Sickle cell anemia is associated with unusual clinical heterogeneity for a Mendelian disorder. Fetal hemoglobin concentration and coincident α thalassemia, both which directly affect the sickle erythrocyte, are the major modulators of the phenotype of disease. Understanding the genetics underlying the heritable subphenotypes of sickle cell anemia would be prognostically useful, could inform personalized therapeutics, and might help the discovery of new "druggable" pathophysiologic targets. Genotype-phenotype association studies have been used to identify novel genetic modifiers. In the future, whole genome sequencing with its promise of discovering hitherto unsuspected variants could add to our understanding of the genetic modifiers of this disease.

Nitric oxide status in sickle cell anemia

In this issue of the Revista Brasileira de Hematologia e Hemoterapia, Elias et al.(1) show the negative correlation between nitrite serum levels and the concentration of fetal hemoglobin (Hb F) in adult patients with sickle cell anemia. Hb F is a predictor of good prognosis in sickle cell anemia suggesting that the nitrite concentration might be used as a biomarker of the disease prognosis(1). Decreased levels of nitrite can indicate a better prognosis as it may be related to an increased production of NO via Hb F(2) . Sickle cell disease (SCD) is an inherited disorder of hemoglobin synthesis. The cause of SCD is a point mutation in the β-globin gene. This genetic abnormality leads to the production of sickle hemoglobin, a protein that has the unique property of polymerizing into long fibers when deoxygenated, thereby decreasing red blood cell deformability and damaging the cell membrane. Sickle cell anemia is characterized by episodes of vaso-occlusion and hemolysis, which are responsible for the clinical manifestations of the disease(3) . This disease is a chronic inflammatory condition that diminishes the reserves of nitric oxide (NO). NO is normally produced by the endothelium and regulates the basal vessel tone, inhibits platelet and hemostatic activation, inhibits adhesion molecules and reduces reactive oxygen species (ROS) levels. The potential importance of NO in SCD has been the subject of much research over the last decade. It is now accepted that the decreased availability of NO is an important mechanism in the pathophysiology of SCD(3). Serum levels of L-arginine, the precursor of NO, are decreased in patients with sickle cell anemia, particularly during vaso-occlusive and acute chest syndrome episodes; these levels are inversely proportional to painful crisis. In the vascular compartment, the hemolytic process causes the release of large amounts of free hemoglobin, heme and arginase. Free plasma hemoglobin, in addition to generating ROS, such as the hydroxyl and superoxide radicals, is also a potent scavenger of nitric oxide. The hydrophobic heme enters the plasma membrane of endothelial cells, releasing iron, which damages the cells causing the release of ROS(3). Arginase, released in plasma L-arginine acts in transforming it into urea and ornithine. The increase in plasma L-ornithine competes with low levels of L-arginine uptake in cellular compartments and may generate pulmonary hypertension and the production of ROS. The half-life of NO in the blood is extremely short because of its rapid reaction with hemoglobin to form methemoglobin and nitrate(4). In conclusion the analysis of the metabolism of NO is important to monitor patients with SCD and biomarkers may be associated with disease progression. This represents the most important advance in understanding the pathogenesis, contributing to a more accurate management of patients(5).

Correlation of low levels of nitrite and high levels of fetal hemoglobin in patients with sickle cell disease at baseline

Background Sickle cell disease is a hemoglobinopathy characterized by hemolytic anemia, increased susceptibility to infections and recurrent vaso-occlusive crises that reduces the quality of life of sufferers. Objective To evaluate the correlation of the levels of lactate dehydrogenase, malonaldehyde and nitrite to fetal hemoglobin in patients with sickle cell disease not under treatment with hydroxyurea in outpatients at a university hospital in Fortaleza, Ceará, Brazil. Methods Forty-four patients diagnosed with sickle cell disease were enrolled at baseline. Diagnosis was confirmed by evaluating the beta globin gene using polymerase chain reaction-restriction fragment length polymorphism. The concentration of fetal hemoglobin was obtained by high-performance liquid chromatography. Serum levels of nitrite, malonaldehyde and lactate dehydrogenase were measured by biochemical methods. Results Significantly higher levels of lactate dehydrogenase, nitrite and malonaldehyde were observed in patients with sickle cell disease compared to a control group. The study of the correlation between fetal hemoglobin levels and these variables showed a negative correlation with nitrite levels. No correlation was found between fetal hemoglobin and malonaldehyde or lactate dehydrogenase. When the study population was stratified according to fetal hemoglobin levels, a decrease in the levels of nitrite was observed with higher levels of fetal hemoglobin (p-value = 0.0415). Conclusion The results show that, similar to fetal hemoglobin levels, the concentration of nitrite can predict the clinical course of the disease, but should not be used alone as a modulator of prognosis in patients with sickle cell disease.

To assess the efficacy of hydroxyurea, in children with homozygous sickle cell disease, in the age group of 1 year to 18 years, at tertiary care hospital

Aims: To evaluate the increase in fetal hemoglobin percentage and MCV in children of sickle cell disease treated with hydroxyurea. To observe the safety of hydroxyurea in treated children. To arrive at the therapeutic dose of the drug to produce the beneficial effects.Settings and Design: A single blind randomized controlled trial of 12 months duration was designed to study the effect of hydroxyurea on sickle cell patients which aims to study the clinical and laboratory effects, in tertiary care hospital.Materials and Methods: Sixty children with severe sickle cell disease in the age group of 1-18 years were studied. Outcomes were measured in terms of decrease in number of vaso-occlusive crises, necessity of blood transfusions, laboratory parameters for different organ functions, blood cell counts and toxicity of hydroxyurea.Results: In patients of sickle, cell disease treated with hydroxyurea there was significant reduction in number and severity of vaso- occlusive crises and necessity of blood transfusions. Total hemoglobin, MCV and fetal hemoglobin percentage were raised significantly. Bone marrow suppression as reflected by the decrease in white blood cell and reticulocyte counts occurred but it was not below the defined toxicity levels. Serum bilirubin was decreased significantly. Conclusions: In the present study, we concluded that, hydroxyurea decreases number of vaso-occlusive crises and need for blood transfusions in children with severe sickle cell disease. It increases total, fetal hemoglobin concentration and MCV. It causes bone marrow suppression thereby decreasing white blood cell and reticulocyte count. However, this fall was never below defined toxicity levels. It causes no hematological, hepatic, renal or general toxicities when given on short-term basis. Overall, hydroxyurea has proved to be safe and efficacious in children between the age groups of 1-18 years.

Цитологічна та біохімічна характеристика периферичної крові щурів за умов експериментального цукрового діабету 1-го типу та дії галеги лікарської

The article contains data on the influence of the Galega officinalis L. extract, on hematological indicators of rats’ peripheral blood under the condition of experimental diabetes mellitus (EDM). It has been shown that the development of EDM is accompanied by increasing glucose concentration, glycated and fetal hemoglobin content, platelet count, and a violation of percentage of different forms of leukocytes. Research conducted by us showed distinct hypoglycemic effect of Galega on the model of streptozotocin diabetes type 1. Admission of the extract to control animals glucose concentration and glycosylated hemoglobin did not differ from control values. Study extract affect glucose metabolism only under its higher concentration, which occurs in diabetes. Admission of Galega officinalis leads to the decrease of fetal and glycated hemoglobin content and platelet count also to the normalization of white blood cell differential count, indicating the activation of protective mechanisms of the body. Keywords: Galega officinalis , diabetes mellitus, peripheral blood, phytotherapy.

Health Care Utilization for Painful Events Is Associated with Early Mortality in a Contemporary Population of Adults with Sickle Cell Anemia

Abstract▪2115▪This icon denotes a clinically relevant abstract Background:Painful vaso-occlusive crises are the most common symptom associated with sickle cell anemia (SCA) and show a large inter-patient variability in the frequency and severity. Historically a high rate of admissions for pain is associated with early mortality in SCA adults. Modern day management of SCA includes screening for life threatening complications such as pulmonary hypertension and disease modifying therapies such as hydroxyurea and chronic blood transfusions. It has also become clear that all acute painful events including some self reported crisis do not lead to ED visit/ hospitalization therefore use of healthcare utilization may underestimate the actual frequency of painful vaso-occlusive crises. The present study determined if ED visit/ hospitalizations for painful vaso-occlusive crises remain a marker of early mortality in the modern era. Methods:To determine the relative contribution of pain and other established risk factors for death in a contemporary adult cohort with sickle cell anemia, we analyzed data of SCA individuals who were evaluated at NIH between February 2001 and June 2007 for screening of the IRB approved protocol to study pulmonary hypertension in sickle cell disease. Self reported history of vaso-occlusive painful events requiring visit to an emergency department (ED) or in-patient hospitalization in 12 months preceding the enrollment was used to stratify study subjects into no-ED visit/ hospitalization or ED visit/hospitalization group. Characteristics between the groups were compared. Doppler echocardiography was used to estimate systolic pulmonary artery pressure through measurement of the tricuspid regurgitation velocity (TRV). Study subjects were followed prospectively and age at death was recorded. Results:One hundred and four (40%) SCA subjects reported no ED visit/ hospitalization while 160 (60%) reported at least one ED visit/ hospitalization in 12 months preceding the enrollment. Although not statistically significant, a higher proportion of study participants in the ED visit/ hospitalization group were on hydroxyurea (38 vs. 69%; P=0.4), and more likely to have received > 10 red cell transfusions (37 vs. 48%; P= 0.09). The no-ED/ hospitalization group and ED/ hospitalization groups were not different in median age (33 vs. 31; P=0.1) or fetal hemoglobin concentration (7.7 vs. 6.9%; P=0.3). Overall forty (15.1%) individuals died during the median follow-up period of 5 years. Cox proportional hazards analysis was performed to determine if severe pain crisis requiring ED visits/hospitalizations were associated with early mortality after adjusting for other known risk factors for mortality (Table 1). Risk for mortality was significantly associated with a history of severe pain crisis requiring ED visit/hospitalization in preceding year (RR 2.81, 95% CI: 1.2–6.4, P= 0.04), elevated TRV ≥ 3 m/s (RR 4.07, 95% CI: 1.3–12.8, P= 0.02) and elevated ferritin (RR 2.31, 95% CI: 1.0–5.0, P= 0.04).Table 1:Cox Proportional Hazards Analysis of MortalityRisk FactorsRR (95% CI)1P2ED visits/hospitalization in past year2.81 (1.2–6.4)0.04TRV, m/sec2.5–2.92.46 (0.8–7.9)0.133.0+4.07 (1.3–12.8)0.02Ferritin2.31 (1.0–5.2)0.041 RR for TRV is given relative to <2.5 m/sec; RR for ED is given relative to zero visits in the past year. RR for ferritin is given for the 75th relative to the 25th percentile.2From Wald χ2 Conclusions:Severe painful episodes requiring health care utilization are associated with premature mortality and despite likely underestimation of actual frequency of pain, remain a marker of disease severity. Elevated TRV a marker of elevated pulmonary arterial pressure and early mortality has a cumulative effect with pain on reduced survival. Consistent with Cooperative Study of Sickle Cell Disease (CSSCD) report, higher hemoglobin (and lower LDH) was associated with reports of ED visit/ hospitalization likely related to increased viscosity. We conclude that as a group, SCA patients reporting severe pain requiring ED visits /hospitalization and elevated TRV are at the highest risk for shortened survival. Disclosures:No relevant conflicts of interest to declare.

Hydroxyurea Treatment of Young Children with Sickle Cell Anemia: Safety and Efficacy of Continued Treatment – the BABY HUG Follow-up Study

Abstract 7▪BABY HUG [Clinical Trials #NCT00006400], an NIH-NICHD sponsored randomized placebo-controlled trial showed that hydroxyurea (HU) administered to 9–18 month old children with sickle cell anemia (SCA) provides substantial clinical benefit. Benefits include a decrease in pain crises, acute chest syndrome events, need for transfusion and hospital admission; hematologic improvement include higher total and fetal hemoglobin concentration, larger red cell size, and lower WBC counts with toxicity limited to transient reduction in absolute neutrophil count (ANC) [Lancet 2011; 377:1663–72]. The parent or guardian of all 176 children who completed at least 18 months of randomized treatment were offered participation in an initial observational BABY HUG Follow-Up Study and 163 (93%) consented to participate. Clinical and laboratory data were collected every 6 months by structured abstraction of the medical record regarding use of clinically prescribed HU (dose escalation recommended), blood counts, clinical imaging, and sickle cell-related events. At the time of enrollment the family did not know their child’s randomized study treatment assignment; 133 (82%) initially chose clinical prescription of open-label HU. Acceptance of HU has remained high through 36 months of follow-up; during each 6 month data collection period 68–75% of participants reported having taken HU.Only 2 patients have left the study (due to relocation) and more than 93% of expected data have been collected. Preliminary analyses as of May 2011, including 417 patient years (pt-yrs) of follow up, demonstrate that in comparison to participants not taking HU, children who continue to take HU have statistically lower rates of pain crises requiring emergency department (ED) visits, episodic transfusions, and hospital admissions for any reason, including acute chest syndrome or febrile illness (see table). The substantial decrease in acute chest syndrome episodes is similar to the effect demonstrated with HU use in the randomized BABY HUG trial in younger infants and consistent with published trials detailing the benefit of HU therapy in older children and adults. The decrease in the rate of admission for febrile events in HU-treated patients is also comparable to that in the randomized trial, but the reason for this benefit is uncertain. There was no difference in hospitalization rates for painful events including dactylitis. Two patients in the non-HU group had a stroke. There were no differences between groups in the frequency of a palpable spleen or rate of acute splenic sequestration crises. Through 36 months of follow up children taking HU had persistently higher hemoglobin and MCV, and lower WBC and ANC than those not taking HU.Event Rate per 100 pt-yrsHUNo HUp valueED visit for Pain Crisis28.853.60.004Episodic Transfusion18.334.00.010Hospital Admission (any cause)74.9133.20.001Acute Chest Syndrome (admission)9.522.30.0001Febrile Illness (admission)30.764.3<.001Pain Crisis (admission)18.630.40.102Results of these analyses including growth and development assessments will enhance our understanding of the impact of HU use in children with SCA starting at a very young age. The accruing data from the BABY HUG Follow-Up Study demonstrate a continuation of the substantial benefits of early HU therapy with no discernable additional toxicities. Ongoing follow up of this cohort is essential to fully define these benefits as children grow, and to observe for late toxicity. Disclosures:Off Label Use: Hydroxyurea is not indicated for treatment of children with sickle cell disease. Use of this medication was for clinical indications and not mandated by this observational study.

The Impact of Hydroxyurea Therapy on the Prevalence of Retinopathy in a Pediatric Sickle Cell Cohort

Abstract 1057 Introduction:Approximately 15–17% of children with HbSS and 11% of those with HbSβ0thalassemia (HbSβ0thal) develop sickle retinopathy (SR) by 18 years of age. Sickle retinopathy is a spectrum of vascular lesions that insidiously evolve as a result of ischemia from vaso-occlusion of capillary and precapillary arterioles in the peripheral retina. These lesions are usually clinically silent but can progress to spontaneous vitreous hemorrhage or retinal detachment and lead to blindness. Therefore, periodic screening retinal exams are recommended for children and adults with sickle cell disease (SCD). Prior reports from Jamaica identified low fetal hemoglobin (HbF) concentration as a potential risk factor for retinopathy in HbSS patients. Hydroxyurea (HU), an increasingly utilized treatment in children with HbSS/HbSB0thal, increases %HbF levels. However, there are no reports describing the effects of HU therapy on the prevalence of retinopathy in children with SCD. In our institution, all children with SCD are referred to ophthalmology for a screening dilated fundoscopic exam (DFE) beginning at 10 years of age and then every 2 years. Objective:Define the prevalence of retinopathy in children with HbSS/HbSβ0thal and analyze the effects of treatment with HU and %HbF levels on the prevalence of retinopathy. Methods:After IRB approval, a retrospective chart review was performed on children with HbSS/HbSβ0thal who received a DFE without fluorescein angiography at St Jude Children’s Research Hospital between 2003 and 2010. Children who received chronic transfusion therapy or hematopoietic stem cell transplantation were excluded from analysis. Records of DFEs were reviewed for a diagnosis of non-proliferative or proliferative SR. Laboratory data were collected at multiple time points from patients’ electronic medical records, including: prior to initiation of HU, following two years of therapy with HU, and immediately prior to diagnosis of retinopathy. Time of observation was defined as birth to diagnosis of SR or to date of last DFE. Summary statistics are reported and the HU and non-HU groups are compared with the Wilcoxon-Mann-Whitney Exact test. Results:Of 182 children referred to ophthalmology, 142 (78%; 126 HbSS, 16 HbSβ0thal) received an initial DFE at a median age of 13.1 ± 3.3 years (range: 1.6–18.7 years). The median number of DFEs per patient was 1.7 ± 1.4 (range: 1–12) during the time of observation. Sickle retinopathy was diagnosed in 9.9% (14/142) of children, at an average age of 15 ± 2.5 years. Of the children who received a DFE, 63% (90/142) had initiated HU therapy at an average age of 9.8 ±4.8 years. The average age at initial DFE was not different in patients receiving HU (13.1 ± 3.2 years) and those who did not (13 ± 3.5 years). The total number of DFEs performed in both groups was also similar (1 ± 1.2 in HU-treated group vs. 1 ± 1.6 in untreated group). Retinopathy was diagnosed in 7.8% (7/90) of children who received HU and 13.5% (7/52) of children who did not (p=0.27). The median age at diagnosis of SR was similar for patients treated with HU (15.4 ± 2.5 years, range: 11.3–18.2 years) compared to untreated children (14.6 ± 2.6 years, range: 10.8–17.3 years; p=0.6). Among patients receiving HU, those who developed retinopathy started treatment at a later age, however the difference was not statistically significant (12.2 ± 5 years vs. 9.3 ± 4.5 years; p=0.23). Among the HU treated group, %HbF following 2 years of HU therapy was similar in those who developed SR (16.1 ± 7.7%) and those who did not (20.0 ± 6.8%; p=0.24). Conclusion:Although the average age at diagnosis with retinopathy in these children with HbSS/HbSβ0thal was similar to previously reported untreated pediatric cohorts, the overall prevalence of retinopathy was lower. Treatment with HU did not affect the development of retinopathy nor did %HbF levels in those treated with HU. More studies are necessary to analyze the effects of HU treatment on development of SR. In the meantime, continued adherence to screening guidelines for ophthalmology evaluations are necessary for children with HbSS and HbSβ0thal being treated with hydroxyurea. Disclosures:Off Label Use: Use of hydroxyurea in children with sickle cell anemia.

Genotype of Beta Thalassemia Patients As Predictors for Prognosis & Management without Blood Transfusion,

Abstract 3199 Introduction:Packed red blood cell (PRC) transfusion with iron chelation, despite their undesirable effects, has been the mainstay of treatment for patients with beta-thalassaemia major. In the recent past introduction of oral medications that augment Hemoglobin F (HbF) has opened new horizons in the management & prognosis of these patients sparing many of them from the hardtimes of blood transfusions & related adversities. Methods:On the basis of our previous studies evaluating the safety & efficacy of Hydroxyurea (HU) in beta thalassemia patients which is an oral HbF augmentation agent, at 10–15 mg/kg/day was used on 238 patients for 24 months under the guidelines of Helsinki’s declaration. Response was measured by using transfusion requirement prior to 6 months period of enrollment in the study as control. Patients were finally divided into 3 groups on the basis of response, Group 1 consisted of Complete Responders, group 2 were partial responders & group 3 were non-responders. Group1 patients needed regular blood transfusions prior to HU therapy while after 24 months they never required transfusion as they maintained their mean Hb at ≥7gm/dL. Partial responders substantially decreased their need for transfusion (less than 50 %), while Non-responders had no decrease in their need for transfusions. All patients’ genetic mutation profiles were investigated upon. Results:Most common genetic mutations observed were IVS1-5 (48%) either homozygous or heterozygous, Fr 8–9 (11.8%), IVS1-1 (10.5%), Cd 30 (7.6 %), Fr 41–42 (6.3%), Cap+1 (3.3%) & Del-619 (8%). Homozygous Xmn-polymorphism was found in 20% & heterozygous Xmn polymorphism was observed in 26% of the patients, while rest 54% had no Xmn polymorphism. Astounding results were observed when these mutations were correlated with response of HU. Response rate was found to be most closely related with gene mutations IVS1-1 (68% Responders), Cd-30 (56% Responders), Cap+1 (38% Responders)IVS1-5 (37% Responders). Xmn polymorphism was found to be significantly associated with Response rate 73% (p-value 0.00). Discussion:It was observed from our study that certain genetic mutations in beta-thalassemia & presence of Xmn polymorohism responds exceptionally well to oral HbF augmentation agents, sparing the patients from the adversities of blood transfusion. Most notably presence of IVS1-1 spared 68% of the patients from blood transfusions while Xmn polymorphism spared 73% of them. It is recommended, therefore, that a new classification be made on the basis of genetic profile of beta thalassemia patients for better treatment & prognosis preventing unnecessary blood transfusions. Disclosures:Ansari:National Institute of Blood Diseases & Bone Marrow Transplantation: Consultancy. Off Label Use: We used Hydroxyurea or hydroxy carbamide. Hydroxycarbamide increases the concentration of fetal hemoglobin. The precise mechanism of action is not yet clear, but it appears that hydroxycarbamide increases nitric oxide levels, causing soluble guanylyl cyclase activation with a resultant rise in cyclic GMP, and the activation of gammaglobin synthesis necessary for fetal hemoglobin. Shamsi:National Institute of Blood Diseases & Bone Marrow Transplantation: Consultancy. Bohray:National Institute of Blood Diseases & Bone Marrow Transplantation: Consultancy. Farzana:National Institute of Blood Diseases: Employment. Erum:National Institute of Blood Diseases: Employment.

Hydroxycarbamide in very young children with sickle-cell anaemia: a multicentre, randomised, controlled trial (BABY HUG)

Sickle-cell anaemia is associated with substantial morbidity from acute complications and organ dysfunction beginning in the first year of life. Hydroxycarbamide substantially reduces episodes of pain and acute chest syndrome, admissions to hospital, and transfusions in adults with sickle-cell anaemia. We assessed the effect of hydroxycarbamide therapy on organ dysfunction and clinical complications, and examined laboratory findings and toxic effects. This randomised trial was undertaken in 13 centres in the USA between October, 2003, and September, 2009. Eligible participants had haemoglobin SS (HbSS) or haemoglobin Sβ(0)thalassaemia, were aged 9-18 months at randomisation, and were not selected for clinical severity. Participants received liquid hydroxycarbamide, 20 mg/kg per day, or placebo for 2 years. Randomisation assignments were generated by the medical coordinating centre by a pre-decided schedule. Identical appearing and tasting formulations were used for hydroxycarbamide and placebo. Patients, caregivers, and coordinating centre staff were masked to treatment allocation. Primary study endpoints were splenic function (qualitative uptake on (99)Tc spleen scan) and renal function (glomerular filtration rate by (99m)Tc-DTPA clearance). Additional assessments included blood counts, fetal haemoglobin concentration, chemistry profiles, spleen function biomarkers, urine osmolality, neurodevelopment, transcranial Doppler ultrasonography, growth, and mutagenicity. Study visits occurred every 2-4 weeks. Analysis was by intention to treat. The trial is registered with ClinicalTrials.gov, number NCT00006400. 96 patients received hydroxycarbamide and 97 placebo, of whom 83 patients in the hydroxycarbamide group and 84 in the placebo group completed the study. Significant differences were not seen between groups for the primary endpoints (19 of 70 patients with decreased spleen function at exit in the hydroxycarbamide group vs 28 of 74 patients in the placebo group, p=0·21; and a difference in the mean increase in DTPA glomerular filtration rate in the hydroxycarbamide group versus the placebo group of 2 mL/min per 1·73 m(2), p=0·84). Hydroxycarbamide significantly decreased pain (177 events in 62 patients vs 375 events in 75 patients in the placebo group, p=0·002) and dactylitis (24 events in 14 patients vs 123 events in 42 patients in the placebo group, p<0·0001), with some evidence for decreased acute chest syndrome, hospitalisation rates, and transfusion. Hydroxyurea increased haemoglobin and fetal haemoglobin, and decreased white blood-cell count. Toxicity was limited to mild-to-moderate neutropenia. On the basis of the safety and efficacy data from this trial, hydroxycarbamide can now be considered for all very young children with sickle-cell anaemia. The US National Heart, Lung, and Blood Institute; and the National Institute of Child Health and Human Development.