André Rolim Belisário

β-globin gene cluster haplotypes in a cohort of 221 children with sickle cell anemia or Sβ⁰-thalassemia and their association with clinical and hematological features.

Background/Aims: βS-Haplotype prevalence and its associations with clinical and hematological characteristics were assessed in Brazilian children with sickle cell anemia or Sβ⁰-thalassemia. Methods: A retrospective randomized cohort study was undertaken with 208 SS and 13 Sβ⁰-thalassemia children derived from the Newborn Screening Program of the state of Minas Gerais. βS-Haplotypes were determined by PCR-RFLP. Results: Thirty-nine percent of the SS subjects had the CAR/CAR genotype, 33% had CAR/Ben, 24% had Ben/Ben, 1% had CAR/Atp, 1% had Ben/Atp, and 1% had Arab-Indian/Ben; 1% could not be characterized. Of the Sβ⁰-thalassemia children, 5 were CAR/undefined, 2 were Ben/undefined, and 1 was CAM/undefined. There was no significant association between βS-haplotypes and the total Hb, Hb F, MCV, MCH, WBC, and reticulocyte count among the SS children. Likewise, no significant association was detected between βS-haplotypes and the frequency of acute chest syndrome episodes, blood transfusions, splenic sequestration, or cerebrovascular disease (high-risk/conditional transcranial Doppler ultrasonography or clinical stroke). A limited number of Sβ⁰-thalassemia children precluded valid analyses. Conclusions: The prevalence of βS-haplotypes in this study is in agreement with the historical records of African slaves brought to the state of Minas Gerais. Furthermore, βS-haplotypes CAR and Ben were not associated with any analyzed feature of children with sickle cell anemia.

Association of alpha-thalassemia, TNF-alpha (-308G>A) and VCAM-1 (c.1238G>C) gene polymorphisms with cerebrovascular disease in a newborn cohort of 411 children with sickle cell anemia

Cerebrovascular disease (CVD) is a severe complication associated with sickle cell anemia. Abnormal transcranial Doppler (TCD) identifies some children at high risk, but other markers would be helpful. This cohort study was aimed at evaluating the effects of genetic biomarkers on the risk of developing CVD in children from Minas Gerais, Brazil. Clinical and hematological data were retrieved from childrens records. Outcomes studied were overt ischemic stroke and CVD (overt ischemic stroke, transient ischemic attack, abnormal TCD, or abnormal cerebral angiography). Out of 411 children, 386 (93.9%) had SS genotype, 23 (5.6%) had Sβ(0)-thal and two had severe Sβ(+)-thal (0.5%). Frequency of CVD was lower in Sβ-thal group (p=0.05). No effect of VCAM-1 polymorphism on stroke or CVD risks was detected. Cumulative incidence of stroke was significantly higher for children with TNF-α A allele (p=0.02) and lower for children with HBA deletion (p=0.02). However, no association between CVD and TNF-α -308G>A was found. CVD cumulative incidence was significantly lower for children with HBA deletion (p=0.004). This study found no association between VCAM1 c.1238G>C and stroke. An association between stroke and TNF-α -308A allele has been suggested. Our results have confirmed the protective role of HBA deletion against stroke and CVD.

Association between ENPP1 K173Q and stroke in a newborn cohort of 395 Brazilian children with sickle cell anemia

To the editor: Transcranial Doppler (TCD) screening and intensification therapy may reduce the risk of stroke from 11% to 1.9% in children with sickle cell anemia (SCA).[1][1] However, the relatively low specificity of TCD screening in identifying individuals at risk of stroke[2][2] highlights the

Glucose-6-Phosphate Dehydrogenase Deficiency in Brazilian Children With Sickle Cell Anemia is not Associated With Clinical Ischemic Stroke or High-Risk Transcranial Doppler.

Stroke is a severe complication of sickle cell anemia (SCA). The role of glucose‐6‐phosphate dehydrogenase (G6PD) deficiency in the development of stroke in children with SCA is controversial.

Clinical, hematological and genetic data of a cohort of children with hemoglobin SD

Introduction The hemoglobin FSD is very uncommon in newborn screening programs for sickle cell disease. In the program of Minas Gerais, Brazil, the clinical course of children with hemoglobin SD was observed to be heterogeneous. The objective of this study was to estimate the incidence (1999–2012) and to describe the natural history of a cohort of newborns with hemoglobin SD. Methods Isoelectric focusing was the primary method used in newborn screening. Polymerase chain reaction-restriction fragment length polymorphism and gene sequencing were used to identify mutant alleles and for haplotyping. Gap-polymerase chain reaction was used to detect alpha-thalassemia. Results Eleven cases of hemoglobin S/D-Punjab and eight of Hb S-Korle Bu were detected. Other variants with hemoglobin D mobility were not identified. All hemoglobin D-Punjab and hemoglobin Korle Bu alleles were associated with haplotype I. Among the children with hemoglobin S/D-Punjab, there were four with the βS CAR haplotype, six with the Benin haplotype, and one atypical. Results of laboratory tests for hemoglobin S/D-Punjab and hemoglobin S-Korle Bu were: hemoglobin 8.0 and 12.3 g/dL (p-value <0.001), leukocyte count 13.9 × 109/L and 10.5 × 109/L (p-value = 0.003), reticulocytes 7.5% and 1.0% (p-value <0.001), hemoglobin F concentration 16.1% and 6.9% (p-value = 0.001) and oxygen saturation 91.9% and 97% (p-value = 0.002), respectively. Only hemoglobin S/D-Punjab children had acute pain crises and needed blood transfusions or hydroxyurea. Those with the Benin βS haplotype had higher total hemoglobin and hemoglobin F concentrations compared to the CAR haplotype. Transcranial Doppler was normal in all children. Conclusion The clinical course and blood cell counts of children with hemoglobin S/D-Punjab were very similar to those of hemoglobin SS children. In contrast, children with hemoglobin S-Korle Bu had clinical course and blood cell counts like children with the sickle cell trait.

Very mild forms of Hb S/beta + -thalassemia in Brazilian children

The clinical phenotype of sickle cell/beta+-thalassemia (Hb S/ +-Thal) is highly variable, and severity is associated with the quantitative degree of decrease in the production of the beta globin chains.1 Evidence shows that differences in the production of hemoglobin A (Hb A) and severity correspond to different molecular beta-thalassemia ( -Thal) mutations.2 A previous report proposed a classification of Hb S/ +-Thal phenotypes based on the relative concentration of Hb A: Type I: 1–7% of Hb A; Type II: 7–14% of Hb A; and Type III: 14–25% of Hb A.2 However, some -Thal mutations lead to low impairment of -globin production and the resulting phenotype does not fit this proposed classification. In a single patient case report, the −92 (C>T) mutation was associated with a high level of Hb A (45%) in an adult Sicilian patient with Hb S/ +-Thal.3 Recently, a combination of two sequence variants, IVS-II839 (T>C) and IVS-II-844 (C>A), was associated with a very mild phenotype of sickle cell disease (SCD).4 Because of the marked clinical variability of Hb S/ +-Thal patients, molecular

De novo alpha 2 hemoglobin gene (HBA2) mutation in a child with hemoglobin M Iwate and symptomatic methemoglobinemia since birth.

Cyanosis in an apparently healthy newborn baby may be caused by hemoglobin variants associated with the formation of methemoglobin, collectively known as M hemoglobins. They should not be confused with genetic alterations in methemoglobin reductase enzyme systems of red cells since treatment and prognosis are completely different. A newborn male child was noted to be significantly cyanotic at birth and is the basis for this report. Hemoglobin isoelectric focusing, acid and alkaline gel electrophoresis, and HBA/HBB gene sequencing were performed for the child, both parents and a sister. The newborn child was treated with methylene blue in an intensive care unit fearing that he had a defective reductase system and exposure to oxidant drugs or toxins. Newborn hemoglobin screening with high performance liquid chromatography was abnormal on the 10th and 45th days but no conclusive diagnosis was reached. Cyanosis persisted up to four years of age with no other symptoms. Hemoglobin M Iwate [alpha2 87(F8) His>Tyr, HBA2:c.262C>T] was detected. It was not present in the childs presumed mother, father, sister, and brother. The analysis of 15 short tandem repeats in the trio demonstrated a de novo mutation occurrence (p-value < 1 × 10−8). The family was reassured that no further action was necessary and genetic counseling was provided. Methemoglobins should be considered for differential diagnosis of cyanosis in newborns even if no familial cases are detected. Except for cosmetic consequences, the clinical course of patients with hemoglobin M Iwate is unremarkable.

The Natural History of Hb S/Hereditary Persistence of Fetal Hemoglobin in 13 Children from the State of Minas Gerais, Brazil

Abstract Children with Hb S (HBB: c.20A > T)/hereditary persistence of fetal hemoglobin (Hb S/HPFH) have a mild clinical phenotype, but some complications have been reported. The natural history of Hb S/HPFH in children from the State of Minas Gerais, Brazil newborn cohort is described. Clinical and hematological data regarding participants’ phenotypes were retrieved from medical records. The HPFH-1, HPFH-2, and HPFH-3 and α-thalassemia (α-thal) deletions were detected by gap-polymerase chain reaction (gap-PCR). Thirteen children were included, nine (69.2%) had the Hb S/HPFH-2 deletion, and four (30.8%) had Hb S/HPFH-1 deletion; 11 children (84.6%) had αα/αα, and two (15.4%) carried the αα/−α3.7 (rightward) deletion. The mean concentration of total hemoglobin (Hb) and Hb F was 12.52 ± 0.56 g/dL and 42.31% ± 1.97%, respectively. Mild microcytosis and hypochromia were observed. We found acute clinical manifestations of sickle cell disease, such as acute chest syndrome (ACS) and acute pain crisis in four children; nine (69.2%) children were completely asymptomatic during the follow-up period. All children were classified as having low-risk transcranial Doppler (TDC). In conclusion, children with Hb S/HPFH have a mild clinical phenotype of sickle cell disease, although acute clinical manifestations may occur. High Hb F levels and absence of anemia are common hematological characteristics.

Glomerular hyperfiltration and β-2 microglobulin as biomarkers of incipient renal dysfunction in cancer survivors

Herein, we aimed to evaluate the occurrence of impaired renal function after cancer treatment with potentially nephrotoxic chemotherapy in children. A cross-sectional study was performed in 41 cancer survivors after chemotherapy with potentially nephrotoxic drugs. 26 (63.4%) children were detected with glomerular hyperfiltration, and urinary levels of β-2 microglobulin (B2MG) were higher than reference range in all patients. Levels of B2MG were positively correlated with plasma creatinine and negatively correlated with glomerular filtration rate. Plasma creatinine, systolic blood pressure and cholesterol were independently associated with B2MG values. The final multivariate model for glomerular hyperfiltration risk included plasma levels of urea and of magnesium. Urinary levels of B2MG and glomerular hyperfiltration may emerge as potential biomarkers of early renal dysfunction in childhood cancer survivors.

Low urinary levels of angiotensin-converting enzyme 2 may contribute to albuminuria in children with sickle cell anaemia

Clinical trials have shown that angiotensin-converting enzyme inhibitors (ACEi) effectively reduce albuminuria in patients with sickle cell anaemia (SCA) (Haymann et al, 2017). However, the utilization of ACEi in SCA was drawn from experience with diabetic nephropathy. No studies have evaluated renin-angiotensin system (RAS) components in patients with SCA, and whether RAS activity plays a role in albuminuria. Animal models of sickle cell disease (SCD) have demonstrated changes in the levels of RAS components (dos Santos et al, 2014; Roy et al, 2018). However, there is currently no data available regarding RAS molecules in humans with SCD, which is crucial to understand the role of RAS components in sickle cell nephropathy, and to identify therapeutic targets. This study sought to evaluate urinary levels of RAS components and the association of these molecules with albuminuria in children with SCA. Detailed methods are provided in Appendix S1. A crosssectional study of 72 children selected from the Minas Gerais state SCA cohort were assigned to two groups: (i) 36 children with SCA who had persistent albuminuria (SCA-PA); (ii) 36 children with SCA with normal albuminuria (SCANA) on hydroxycarbamide therapy, sex and age-matched with PA group. The study protocol was approved by our Ethics Committee. Medical records were reviewed to obtain demographic, clinical, laboratory and management data. Random spot urine specimens were collected during routine visits. Albumin/creatinine ratio and total urinary protein