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The clinical and phenotypic heterogeneity of patients with sickle cell anemia (SCA) is influenced by environmental and genetic factors. Several genetic modifiers, such as the KLOTHO (KL) gene, have been associated with SCA clinical outcomes. The KL gene and its encoded proteins are implicated in important biological pathways, which affect the disease’s pathophysiology, such as expression of adhesion molecules VCAM-1 and ICAM-1, oxidative stress, and nitric oxide biology. Here, we evaluated the clinical relevance of two polymorphisms found on the KL gene (rs685417 and rs211239) in 588 unrelated patients with SCA. Genotyping analyses were performed using the TaqMan system. The KL rs211239 was associated with increased number of vaso-occlusive crisis (VOCs) per year (P = 0.001), while KL rs685417 was associated with increased frequency of stroke (P = 0.034), priapism (P = 0.011), number of complications (P = 0.019), and with a lower incidence of priapism (P = 0.036). Additionally, the associations with VOCs, stroke, and priapism remained consistent in multivariate analyses (P < 0.05). Our data highlight the clinical importance of KL in SCA.

Alpha thalassemia and beta-globin haplotype are considered classical genetic disease modifiers in sickle cell anemia (SCA) causing clinical heterogeneity. Nevertheless, their functional impact on SCA disease emergence and progression remains elusive. To better understand the role of alpha thalassemia and beta-globin haplotype in SCA, we performed a retrospective study evaluating the clinical manifestations of 614 patients. The univariate analysis showed that the presence of alpha-thalassemia -3.7-kb mutation (αα/-α and -α/-α) decreased the risk of stroke development (p = 0.046), priapism (p = 0.033), and cholelithiasis (p = 0.021). Furthermore, the cumulative incidence of stroke (p = 0.023) and cholelithiasis (p = 0.006) was also significantly lower for patients carrying the alpha thalassemia -3.7-kb mutation. No clinical effects were associated with the beta-globin haplotype analysis, which could be explained by the relatively homogeneous haplotype composition in our cohort. Our results reinforce that alpha thalassemia can provide protective functions against hemolysis-related symptoms in SCA. Although, several genetic modifiers can impact the inflammatory state of SCA patients, the alpha thalassemia mutation remains one of the most recurrent genetic aberration and should therefore always be considered first.

Hyperbilirubinemia in patients with sickle cell anemia (SCA) as a result of enhanced erythrocyte destruction, lead to cholelithiasis development in a subset of patients. Evidence suggests that hyperbilirubinemia may be related to genetic variations, such as the UGT1A1 gene promoter polymorphism, which causes Gilbert syndrome (GS). Here, we aimed to determine the frequencies of UGT1A1 promoter alleles, alpha thalassemia, and βS haplotypes and analyze their association with cholelithiasis and bilirubin levels. The UGT1A1 alleles, -3.7 kb alpha thalassemia deletion and βS haplotypes were determined using DNA sequencing and PCR-based assays in 913 patients with SCA. The mean of total and unconjugated bilirubin and the frequency of cholelithiasis in GS patients were higher when compared to those without this condition, regardless of age (P 0.05). However, not cholelithiasis but total and unconjugated bilirubin levels were associated with βS haplotype. These findings confirm in a large cohort that the UGT1A1 polymorphism influences cholelithiasis and hyperbilirubinemia in SCA. HbF and alpha thalassemia also appear as modulators for cholelithiasis risk.

Alpha thalassemia and beta-globin haplotype are considered classical genetic disease modifiers in sickle cell anemia (SCA) causing clinical heterogeneity. Nevertheless, their functional impact on SCA disease emergence and progression remains elusive. To better understand the role of alpha thalassemia and beta-globin haplotype in SCA, we performed a retrospective study evaluating the clinical manifestations of 614 patients. The univariate analysis showed that the presence of alpha-thalassemia -3.7-kb mutation (αα/-α and -α/-α) decreased the risk of stroke development (p = 0.046), priapism (p = 0.033), and cholelithiasis (p = 0.021). Furthermore, the cumulative incidence of stroke (p = 0.023) and cholelithiasis (p = 0.006) was also significantly lower for patients carrying the alpha thalassemia -3.7-kb mutation. No clinical effects were associated with the beta-globin haplotype analysis, which could be explained by the relatively homogeneous haplotype composition in our cohort. Our results reinforce that alpha thalassemia can provide protective functions against hemolysis-related symptoms in SCA. Although, several genetic modifiers can impact the inflammatory state of SCA patients, the alpha thalassemia mutation remains one of the most recurrent genetic aberration and should therefore always be considered first.

Objective: This study aimed to evaluate whether the single nucleotide polymorphisms (SNPs) +191 C>A (rs4644) and +292 A>C (rs4652) of the LGALS3 gene and the serum levels of galectin-3 (gal-3) are associated with clinical events in patients with sickle cell anemia (SCA). Methods: SNP +191 and +292 of the LGALS3 gene were detected by the TaqMan PCR system in real time. Gal-3 levels were measured in serum by ELISA. The study included 322 patients, mean age 36 (21-84). Results: AA and CA genotypes of the +191 region were related to lower levels of gal-3 when compared to CC genotype (p=0.0296). Lower level of gal-3 was also associated with the +191/+292 (AA/CC; CA/CC) diplotypes (p=0.0137) compared to the diplotypes (CC/AA; CC/CC; CC/AC; CA/AC). There was no association between serum levels of galectin-3 and genotype frequencies of the LGALS3 +191 and +292 polymorphisms with clinical events in SCA. Conclusion: The polymorphisms +191 and +292 of the LGALS3 are associated to decrease in serum levels of gal-3. However, no association of polymorphisms and levels of gal-3 with clinical events was observed in patients SCA. Objetivo: Este estudio tuvo como objetivo evaluar si los polimorfismos de un solo nucleótido (SNPs) +191 C>A (rs4644) y +292 A>C (rs4652) del gen LGALS3 y los niveles séricos de galectina-3 (gal-3) están asociados con los eventos clínicos de pacientes con anemia falciforme (AF). Métodos: SNPs +191 y +292 en LGALS3 se estudiaron utilizando el sistema de PCR en tiempo real TaqMan; Los niveles séricos de gal-3 se midieron por ELISA. El estudio incluyó a 322 pacientes con AF, edad media 36 (21-84). Resultados: los genotipos AA y CA de la región +191 se relacionaron con niveles más bajos de gal-3 en comparación con el genotipo CC (p = 0.0296). El nivel más bajo de gal-3 también se asoció con los diplotipos + 191 / + 292 (AA / CC; CA / CC) (p = 0.0137) en comparación con los diplotipos (CC/AA; CC/CC; CC/AC; CA/CA). No hubo asociación entre los niveles séricos de galectina-3 y las frecuencias de genotipo de los polimorfismos LGALS3 +191 y +292 con eventos clínicos en SCA. Conclusión: Los polimorfismos +191 y +292 de LGALS3 están asociados con una disminución en los niveles séricos de gal-3. Sin embargo, no se observó asociación de polimorfismos y niveles séricos de gal-3 con eventos clínicos en pacientes con AF. Objetivo: Este estudo teve como objetivo avaliar se os polimorfismos de nucleotídeo único (SNPs) +191 C>A (rs4644) e +292 A>C (rs4652) do gene LGALS3 e os níveis séricos da galectina-3 (gal-3) estão associados com os eventos clínicos de pacientes com anemia falciforme (AF). Métodos: Os polimorfismos +191 e +292 no LGALS3 foram estudados usando o sistema de PCR em tempo real, pela metodologia TaqMan; os níveis séricos da gal-3 foram medidos pela técnica de ELISA. O estudo incluiu 322 pacientes com AF, média de idade 36 (21-84). Resultados: Os genótipos AA e CA da região +191 foram relacionados a níveis mais baixos de gal-3 quando comparados ao genótipo CC (p = 0,0296). Níveis mais baixos de gal-3 também foram associados aos diplótipos +191/+292 (AA/CC; CA/CC) (p = 0,0137) em comparação com os diplótipos (CC/AA; CC/CC; CC/AC; CA/AC). Não houve associação entre os níveis séricos da gal-3 e as frequências genotípicas dos SNPs LGALS3 +191 e +292 com os eventos clínicos na AF. Conclusão: Os SNPs +191 e +292 do LGALS3 estão associados à diminuição dos níveis séricos de gal-3. No entanto, não foi observada associação de polimorfismos e níveis séricos da gal-3 com os eventos clínicos em pacientes com AF.

Objective: This study has as objective to verify whether MBL2 gene polymorphisms are related to the occurrence of cerebrovascular disease (CD) in sickle cell anemia (SCA) patients. Methods: Overall, 259 unrelated SCA patients were enrolled. The patients were divided into three groups: control group, stroke group ad range of risk group. Peripheral blood samples were collected and DNA extraction was performed. All patients were genotyped for exon 1, promoter region -221 and promoter region -550 of MBL2 gene, along with β-globin gene haplotypes. Results: Concerning the genotyping of the MBL2, there was no difference in the frequency of allelic and genotypic variants of the exon 1 and the promoter regions -221 and -550 of the MBL2 gene among the studied groups. Conclusion: Despite the small number of patients, and the lack of association between MBL2 polymorphisms and CD, our study represents an effort to understand the impact of MBL2 polymorphisms in the clinical outcome of patients with SCA. Objetivo: Este estudio tiene como objetivo verificar si los polimorfismos del gen MBL2 están relacionados con la aparición de enfermedad cerebrovascular (EC) en pacientes con anemia falciforme (AF). Métodos: en total, se incluyeron 259 pacientes con AF no relacionada. Los pacientes se dividieron en tres grupos: grupo de control, grupo de accidente cerebrovascular y rango de riesgo. Se recogieron muestras de sangre periférica y se realizó extracción de DNA. Todos los pacientes fueron genotipados para el exón 1, la región promotora -221 y la región promotora -550 del gen MBL2, junto con los haplotipos del gen de la β-globina. Resultados: con respecto al genotipo MBL2, no hubo diferencia en la frecuencia de variantes alélicas y genotípicas del exón 1 y en las regiones promotoras -221 y -550 del gen MBL2 entre los grupos estudiados. Conclusión: a pesar del pequeño número de pacientes y la falta de asociación entre los polimorfismos MBL2 y EC, nuestro estudio representa un esfuerzo por comprender el impacto de los polimorfismos MBL2 en el curso clínico de los pacientes com AF. Objetivo: Este estudo tem como objetivo verificar se os polimorfismos do gene MBL2 estão relacionados com a ocorrência de doença cerebrovascular (DC) em pacientes com anemia falciforme (AF). Métodos: No total, 259 pacientes com AF não relacionados foram incluídos. Os pacientes foram divididos em três grupos: grupo controle, grupo acidente vascular cerebral (AVC) e faixa de risco. Amostras de sangue periférico foram coletadas e foi realizada extração de DNA. Todos os pacientes foram genotipados para o éxon 1, região promotora -221 e região promotora -550 do gene MBL2, juntamente com os haplótipos do gene da β-globina. Resultados: Em relação à genotipagem do MBL2, não houve diferença na frequência das variantes alélicas e genotípicas do éxon 1 e nas regiões promotoras -221 e -550 do gene MBL2 entre os grupos estudados. Conclusão: Apesar do pequeno número de pacientes e da falta de associação entre polimorfismos do MBL2 e DC, nosso estudo representa um esforço para entender o impacto dos polimorfismos do MBL2 no curso clínico de pacientes com AF.

Sickle cell anemia (SCA) presents heterogenous clinical manifestations that cannot be explained solely by alterations to hemoglobin (Hb); other components such as endothelial adhesion, thrombosis and inflammation may be involved. The mannose-binding lectin (MBL) has an important role in innate immunity and inflammatory diseases. In this report, we describe an association between MBL2 polymorphism related to low production of serum MBL and the frequency of vasoocclusive events (FVOE) in children ≤ 5 years old with SCA (p = 0.0229; OR 5.55; CI 1.11-27.66). Further studies are needed to explore the role of low MBL2 in the pathophysiology of vasoocclusive events in SCA.

Portal vein thrombosis is considered a vaso-occlusive process that can appear during the course of hepatosplenic Schistosoma mansoni, but may result from impaired portal blood flow or be associated with acquired or inherited thrombophilic factors. Here, we report the case of a 67-year-old woman who developed thrombocytopenia as a result of hypersplenism. Following the diagnosis of hepatosplenic schistosomiasis, portal vein thrombosis was detected by ultrasound examination, while haematological tests revealed low levels of protein C (43.3%) and high levels of factor VIII (183.1%). The pathogenesis of portal vein thrombosis remains unclear in some patients with S. mansoni. We recommend, therefore, that early clinical and haemostatic investigations are done to evaluate risk of portal vein thrombosis and hence avoid further complications.

The aim of this study was to investigate the association between three polymorphisms involved in the oxidative stress pathway and fetal hemoglobin (Hb F) levels in patients with sickle cell anemia in a Brazilian population. One hundred and seven patients with sickle cell anemia were recruited for genomic DNA extraction. The levels of Hb F, sex and age were evaluated. Three polymorphisms, rs4673:T>C and rs9932581:G>A in the CYBA gene and rs2071746:A>T in the HMOX1 gene, were identified through direct sequencing. Hb F levels were not associated with sex, age, or the polymorphisms rs4673:T>C and rs9932581:G>A. However, the TT genotype of the rs2071746:A>T polymorphism was associated with increased levels of Hb F (p value = 0.0131). We observed an association between the TT genotype of the rs2071746:A>T polymorphism, present in the HMOX1 gene, and increased levels of Hb F, indicating the presence of a new marker related to Hb F levels in sickle cell anemia patients.

Background. Previous studies on the role of inflammation in the pathophysiology of sickle cell disease (SCD) suggested that theCCR5Δ32allele, which is responsible for the production of truncated C-C chemokine receptor type 5 (CCR5), could confer a selective advantage on patients with SCD because it leads to a less efficient Th1 response. We determined the frequency of theCCR5Δ32polymorphism in 795 Afro-Brazilian SCD patients followed up at the Pernambuco Hematology and Hemotherapy Center, in Northeastern Brazil, divided into a pediatric group (3 months–17 years,n=483) and an adult group (18–70 years,n=312). The adult patients were also compared to a healthy control group (blood donors, 18–61 years,n=247).Methods. TheCCR5/CCR5Δ32polymorphism was determined by allele-specific PCR.Results. No homozygous patient for theCCR5Δ32allele was detected. The frequency of heterozygotes in the study population (patients and controls) was 5.8%, in the total SCD patients 5.1%, in the children 5.4%, in the adults with SCD 4.8%, and in the adult controls 8.1%. These differences did not reach statistical significance.Conclusions. Our findings failed to demonstrate an important role of theCCR5Δ32allele in the population sample studied here.

The hereditary deficiency of antioxidant enzymes when associated with sickle cell anemia (SCA) further contributes to the oxidation of hemoglobin S, which increases the formation of degradation products of this hemoglobin. The glutathione S transferases play an important role in the conjugation of glutathione to endogenous products of peroxidation of lipids and protect cells from the deleterious effects of oxidative stress. We analyzed genomic DNA from 278 patients with sickle cell anemia to correlate the genotypes GSTT1 and/or GSTM1 null (determined by multiplex PCR technique) and the clinical manifestations of the disease. 27% of patients showed absence of the GSTM1 gene and 15% had absence of GSTT1. The GSTM1 and GSTT1 null genotypes were found in 11% of the population. The risk of individuals with the GSTT1 null genotype developing acute chest syndrome and aseptic necrosis of the femoral head were, respectively, 10 and 6.3 times higher when compared with those individuals who had of this gene. Patients with GSTM1 null showed a risk 3.9 times higher to develop stroke and high risk for malleolar ulcers and acute chest syndrome (OR = 6.9 and 4.2, respectively). The individuals with the GSTM1 and GSTT1 null genotypes showed a higher chance of developing acute chest syndrome, malleolar ulcer and aseptic necrosis of the femoral head. The absence of GSTT1 and/or GSTM1 was an important risk factor for increasing the morbidity of SCA, especially in regard to acute chest syndrome.

Vaso-occlusion is a determinant for most signs and symptoms of sickle-cell anemia (SCA). The mechanisms involved in the pathogenesis of vascular complications in SCA remain unclear. It is known that genetic polymorphisms associated with thrombophilia may be potential modifiers of clinical features of SCA. The genetic polymorphisms C677T and A1298C relating to the enzyme methylenetetrahydrofolate reductase (MTHFR), a clotting Factor V Leiden mutation (1691G→A substitution of Factor V Leiden), and the mutant prothrombin 20210A allele were analyzed in this study. The aim was to find possible correlations with vascular complications and thrombophilia markers in a group of SCA patients in Pernambuco, Brazil. The study included 277 SCA patients, divided into two groups: one consisting of 177 nonconsanguineous SCA patients who presented vascular manifestations of stroke, avascular necrosis, leg ulcers, priapism, and acute chest syndrome (group 1); and the other consisting of 100 SCA patients without any reported vascular complication (group 2). Molecular tests were done using either polymerase chain reaction (PCR) restriction fragment length polymorphism or allele-specific PCR techniques. Comparisons between the groups were made using the χ(2) test. The 677 CT and TT genotypes showed a significant risk of vascular complications (p=0.015). No significant associations between the groups were found when samples were analyzed for the MTHFR A1298C allele (p=0.913), Factor V G1691 (p=0.555), or prothrombin G20210A mutation (p=1.000). The polymorphism MTHFR C677T seemed to be possibly predictive for the development of some vascular complications in SCA patients among this population.

The JAK2 V617F mutation is associated with three myeloproliferative neoplasms (MPNs): polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). It generates an unregulated clonal hematopoietic progenitor and leads to abnormal increased proliferation of one or more myeloid lineages. Subjects bearing this mutation may present more frequently with complications such as thrombosis and bleeding, and no specific treatment has yet been developed for BCR-ABL-negative JAK2 V617F-negative MPNs.To determine the prevalence of JAK2 V617F in MPNs in Pernambuco, Brazil, and to compare it with previous studies.144 blood samples were collected at the Hospital of Hematology of the HEMOPE Foundation and were genotyped by polymerase chain reaction-restriction fragment length polymorphism with BsaXI enzymatic digestion.88% (46/52) of the patients with PV, 47% (39/81) with ET, and 77% (8/11) with PMF were positive for JAK2 V617F, while more than 35% of the individuals were JAK2 V617F-negative, confirming a high prevalence of this abnormality in MPNs, more frequently with a low mutated allele burden, similar to what has been reported in other Western countries, despite differences among methods used to detect this mutation. Screening for JAK2 V617F may allow specific management of these diseases with JAK2 inhibitors in the future and highlights the need for further studies on the pathogenesis of BCR-ABL-negative JAK2 V617F-negative MPNs.