Persistence Of Fetal Hemoglobin Research Articles

Title: Single-Cell Transcriptome Analyses Further Elucidate the Mechanism of Action of Eed Inhibition on HbF Induction

Mutations in the gene encoding the hemoglobin β (HBβ) subunit lead to red blood cell disorders such as Sickle Cell Disease (SCD) and β-thalassemia. Hemoglobin γ subunit, a fetal counterpart of HBβ, can relieve the disease symptoms by forming a functional complex with hemoglobin a subunit, thereby compensating for the mutated HBβ. The expression of β-like hemoglobins is tightly regulated during development, with a switch from the fetal ortholog (γ) to the adult ortholog (β) shortly after birth. Maintenance of HBγ is therapeutically beneficial for SCD patients and is observed clinically in individuals with mutations for both SCD and hereditary persistent fetal hemoglobin (HPFH) who exhibit less severe or even no SCD-related symptoms. Polycomb repressive complex (PRC) proteins have been shown to regulate the expression of β-like hemoglobins. Our previous research has demonstrated inhibitors targeting the PRC2 protein embryonic ectoderm development (EED) upregulate fetal hemoglobin (HbF) expression, leading to the formation of HbF complexes in both human CD34 + HSC-derived erythroid cells and the Townes SCD mouse model. However, the functional mechanisms of PRC complexes in this regulatory process and their target genes remain to be clarified. In our study, we conducted single-cell transcriptome analyses on erythroid cells derived from CD34 + HSCs collected at various stages of differentiation. We identified several erythroid subpopulations, from early progenitors to maturing cells, within the CD34 + derived erythroid cell cultures. HbF + and HbF - cell populations were also identified using known markers. To explore the mechanisms underlying HbF upregulation by EED inhibition, we compared single-cell transcriptome profiles of samples treated with and without an EED inhibitor. Our findings reveal that EED inhibitor treatment consistently leads to increased HbF + populations across multiple donors without impacting erythroid cell differentiation. Differential gene expression analysis revealed sequential changes in the expression of previously identified HbF regulators during differentiation in the presence of EED inhibitor treatment. Through pseudotime analyses, we identified erythroid progenitor cells undergoing fate bifurcation, leading to an increase in the lineage of mature cells that express HBγ. Finally, by comparing the pseudotime trajectories of different samples, we identified genes whose expression changed along the differentiation trajectories in response to EED inhibitor treatment which influence cell fate. Overall, this study provides a deeper understanding of the relevant biological pathways that respond to EED inhibition and potential for the identification of novel molecular targets for HbF induction.

Molecular Epidemiological Characteristics and Differential Diagnosis of Common δβ-Thalassemia/HPFH

To investigate the molecular epidemiological characteristics of common δβ-thalassemia/hereditary persistence of fetal hemoglobin(HPFH) in the prepregnant population in Huadu, and to provide a laboratory basis for prevention and control of thalassemia. Blood samples of childbearing age people in Huadu District of Guangzhou who participated in free thalassemia testing from January 2016 to July 2021 were collected for hematological parameters analysis and hemoglobin electrophoresis. Chinese Gγ+(Aγδβ)0-thalassemia, SEA-HPFH and Taiwanese deletion β-thalassemia were detected by Gap-PCR in the samples with higher HbF(≥5%). Primers were designed for the proximal HBG1 and HBG2 promoter, and the point mutations in the proximal promoter region were detected by Sanger sequencing. Hematology parameters data were statistically analyzed. Among 27 088 samples, Thirteen cases of Chinese Gγ+(Aγδβ)0-thalassemia and thirty-three cases of SEA-HPFH were detected, which including 3 cases of Chinese Gγ+(Aγδβ)0/βN compounded with --SEA/αα and three cases of SEA-HPFH/βN compounded with --SEA/αα. 6 carriers with Aγ-196 C>T were also detected; No Taiwanese thalassemia genetype was detected. The total detection rate of common δβ-thalassemia/HPFH was 0.19% (52/27 088). There were significant differences in the levels of MCV, MCH, HbA2, and HbF among Chinese Gγ+(Aγδβ)0-thalassemia, SEA-HPFH, Aγ-196 C>T (P<0.001). The hematological parameters of Aγ-196C>T combined with α0-thalassemia were similar to those of Chinese Gγ+(Aγδβ)0-thalassemia carriers, and only HbA2 was significantly lower than that of the latter, which was helpful for clinical identification. δβ-thalassemia/HPFH should be included in the scope of thalassemia prevention program in the prepregnant population in Huadu District, and hematological parameters can provide some basis for identifying different types of δβ-thalassemia/HPFH.

In Through the Out Door: Editas' CEOs Grasp Opportunities Amidst Challenges

In Through the Out Door: Editas' CEOs Grasp Opportunities Amidst Challenges

Phenotypic variation in sickle cell disease: the role of beta globin haplotype, alpha thalassaemia and fetal haemoglobin in HbSS

Introduction The haematological and clinical feature vary markedly between the different genotypes of sickle cell disease. Even within the single genotype of homozygous sickle cell disease (HbSS), there is marked variability which is presumed to result from interacting genetic and environmental factors. Areas Covered The classification of the different genotypes of sickle cell disease with approximate prevalence at birth in different communities and some of the major clinical and haematological differences. This assessment includes three potential genetic factors influencing haematology and clinical outcome in HbSS, the beta globin haplotype, alpha thalassaemia and persistence of fetal haemoglobin (HbF). Expert Opinion The author is a clinician with experience of sickle cell disease primarily in Jamaica but also in Greece, Uganda, Saudi Arabia and India. It is therefore necessarily an account of clinical data and does not address current debates on molecular mechanisms. Most data derive from Jamaica where efforts have been made to reduce any symptomatic bias by long term follow-up of patients all over the island and further reduced by a cohort study based on newborn screening which has been in operation for over 48 years.

Epigenomic analysis of KLF1 haploinsufficiency in primary human erythroblasts

Haploinsufficiency for the erythroid-specific transcription factor KLF1 is associated with hereditary persistence of fetal hemoglobin (HPFH). Increased HbF ameliorates the symptoms of β-hemoglobinopathies and downregulation of KLF1 activity has been proposed as a potential therapeutic strategy. However, the feasibility of this approach has been challenged by the observation that KLF1 haploinsufficient individuals with the same KLF1 variant, within the same family, display a wide range of HbF levels. This phenotypic variability is not readily explained by co-inheritance of known HbF-modulating variants in the HBB, HBS1L-MYB and/or BCL11A loci. We studied cultured erythroid progenitors obtained from Maltese individuals in which KLF1 p.K288X carriers display HbF levels ranging between 1.3 and 12.3% of total Hb. Using a combination of gene expression analysis, chromatin accessibility assays and promoter activity tests we find that variation in expression of the wildtype KLF1 allele may explain a significant part of the variability in HbF levels observed in KLF1 haploinsufficiency. Our results have general bearing on the variable penetrance of haploinsufficiency phenotypes and on conflicting interpretations of pathogenicity of variants in other transcriptional regulators such as EP300, GATA2 and RUNX1.

Novel variants in Krueppel like factor 1 that cause persistence of fetal hemoglobin in In(Lu) individuals

Beta-hemoglobinopathies become prominent after birth due to a switch from γ-globin to the mutated β-globin. Haploinsufficiency for the erythroid specific indispensable transcription factor Krueppel-like factor 1 (KLF1) is associated with high persistence of fetal hemoglobin (HPFH). The In(Lu) phenotype, characterized by low to undetectable Lutheran blood group expression is caused by mutations within KLF1 gene. Here we screened a blood donor cohort of 55 Lutheran weak or negative donors for KLF1 variants and evaluated their effect on KLF1 target gene expression. To discriminate between weak and negative Lutheran expression, a flow cytometry (FCM) assay was developed to detect Lu antigen expression. The Lu(a−b−) (negative) donor group, showing a significant decreased CD44 (Indian blood group) expression, also showed increased HbF and HbA2 levels, with one individual expressing HbF as high as 5%. KLF1 exons and promoter sequencing revealed variants in 80% of the Lutheran negative donors. Thirteen different variants plus one high frequency SNP (c.304 T > C) were identified of which 6 were novel. In primary erythroblasts, knockdown of endogenous KLF1 resulted in decreased CD44, Lu and increased HbF expression, while KLF1 over-expressing cells were comparable to wild type (WT). In line with the pleiotropic effects of KLF1 during erythropoiesis, distinct KLF1 mutants expressed in erythroblasts display different abilities to rescue CD44 and Lu expression and/or to affect fetal (HbF) or adult (HbA) hemoglobin expression. With this study we identified novel KLF1 variants to be include into blood group typing analysis. In addition, we provide further insights into the regulation of genes by KLF1.

Characterization of β-Globin Gene Cluster Deletions Using Multiplex-Gap Polymerase Chain Reaction (PCR) and Multiplex Ligation-Dependent Probe Amplification (MLPA)

Background : Deletions in the β-globin gene cluster are usually rare, problematic to detect, and subsequently possess a challenge in many diagnostic laboratories. They are normally related to the heterozygous of the delta beta (δβ) thalassemia, hereditary persistence of fetalhemoglobin (HPFH) and some of the hemoglobin variants. These disorders are typically presented by elevated levels of hemoglobin F (Hb F), but with low to normal hemoglobin A2 (Hb A2). However, despite their existence, there is still a limited number of studies focusing on this area, hence no definitive diagnosis could be conclusively established. Therefore, this pilot study was carried out to fill this knowledge gap.&#x0D; Methods: In this study, screening of the selected deletional mutations in the β-globin gene cluster among patients with Hb F (&gt;1%) and Hb A2 (&lt;4%) were performed using multiplex Gap-PCR and multiplex ligation-dependent probe amplification (MLPA).&#x0D; Results: The results showed that out of 54 samples tested using multiplex Gap-PCR against four target deletions; Thai (δβ)°-thalassemia, HPFH-6, Siriraj J and HbLepore, one sample was found positive with Thai (δβ)°-thalassemia. Further findings from the MLPA screening on 12 randomly selected samples revealed that another patient was positive with double deletions within the region of the β-globin gene cluster. These deletions occur at the gamma-globin gene 1 (HBG1) and gamma-globin gene 2 (HBG2) in exon 3.&#x0D; Conclusions: In conclusion, this study highlighted the importance of these deletions’ characterization using multiplex Gap-PCR and MLPA which helps in establishing a definitive diagnosis among this selected group of patients.&#x0D; Bangladesh Journal of Medical Science Vol.20(3) 2021 p.618-624

Mild dyserythropoiesis and \u03b2-like globin gene expression imbalance due to the loss of histone chaperone ASF1B

The expression of the human β-like globin genes follows a well-orchestrated developmental pattern, undergoing two essential switches, the first one during the first weeks of gestation (ε to γ), and the second one during the perinatal period (γ to β). The γ- to β-globin gene switching mechanism includes suppression of fetal (γ-globin, HbF) and activation of adult (β-globin, HbA) globin gene transcription. In hereditary persistence of fetal hemoglobin (HPFH), the γ-globin suppression mechanism is impaired leaving these individuals with unusual elevated levels of fetal hemoglobin (HbF) in adulthood. Recently, the transcription factors KLF1 and BCL11A have been established as master regulators of the γ- to β-globin switch. Previously, a genomic variant in the KLF1 gene, identified by linkage analysis performed on twenty-seven members of a Maltese family, was found to be associated with HPFH. However, variation in the levels of HbF among family members, and those from other reported families carrying genetic variants in KLF1, suggests additional contributors to globin switching. ASF1B was downregulated in the family members with HPFH. Here, we investigate the role of ASF1B in γ- to β-globin switching and erythropoiesis in vivo. Mouse-human interspecies ASF1B protein identity is 91.6%. By means of knockdown functional assays in human primary erythroid cultures and analysis of the erythroid lineage in Asf1b knockout mice, we provide evidence that ASF1B is a novel contributor to steady-state erythroid differentiation, and while its loss affects the balance of globin expression, it has no major role in hemoglobin switching.

Genetic research and clinical analysis of deletional Chinese G\u03b3+(A\u03b3\u03b4\u03b2)0 -thalassemia and Southeast Asian HPFH in South China

Chinese Gγ+(Aγδβ)0-thalassemia and SEA-HPFH are the most common types of β-globin gene cluster deletion in Chinese population. The aim of the study was to analyze clinical features of deletional Chinese Gγ+(Aγδβ)0-thalassemia and Southeast Asian hereditary persistence of fetal hemoglobin (SEA-HPFH) in South China. A total of 930 subjects with fetal hemoglobin (HbF) level ≥ 2% were selected on genetic research of Chinese Gγ+(Aγδβ)0-thalassemia and SEA-HPFH. The gap polymerase chain reaction was performed to identify the deletions. One hundred cases of Chinese Gγ+(Aγδβ)0-thalassemia were detected, including 90 cases of Chinese Gγ+(Aγδβ)0/βN-thalassemia, 7 cases of Chinese Gγ+(Aγδβ)0 /βN-thalassemia combined with α-thalassemia, 2 cases of Chinese Gγ+(Aγδβ)0-thalassemia combined with β-thalassemia, and 1 case of Chinese Gγ+(Aγδβ)0-thalassemia combined with β-gene mutation. One hundred nine cases of SEA-HPFH were detected, including 97 cases of SEA-HPFH/βN, 9 cases of SEA-HPFH/βN combined with α-thalassemia, 2 cases of SEA-HPFH combined with β-thalassemia, and 1 case of SEA-HPFH combined with β-gene mutation. Statistical analysis indicates significant differences in MCV (mean corpuscular volume), MCH (mean corpuscular hemoglobin), and HbA2 and HbF levels between Chinese Gγ+(Aγδβ)0-thalassemia heterozygotes and SEA-HPFH heterozygotes (P < 0.001). There are statistical differences in hematological parameters between them. Clinical phenotypic analysis can provide guidance for genetic counseling and prenatal diagnosis.

The Effect of Five Single Nucleotide Polymorphisms on Hb F Variation of β-Thalassemia Traits and Hematologically Normal Individuals in Southeast Turkey

β-Thalassemia (β-thal) is caused by deficiency of β-globin chain synthesis and leads to the accumulation of unstable globin chain production. This results in a higher Hb F level in order to neutralize the excess α chains. In addition, γ-globin gene expression, due to genetic factors after birth, leads to increased Hb F levels in adulthood [hereditary persistence of fetal hemoglobin (Hb) (HPFH)]. In this study, the relationship between β-thal trait and individuals with suspected HPFH and a control group was investigated in Adıyaman, Turkey. Single nucleotide polymorphism (SNP) analyses were performed in five different polymorphic regions using real-time polymerase chain reaction (qPCR) methods [rs4671393 (G>A), rs766432 (A>C), rs9402686 (G>A), rs28384513 (T>G), rs1609812 (A>G)]. No significant difference was found between the control and β-thal group in the codominant inheritance model in the rs1609812 (A>G) polymorphism region only, while all the other polymorphic regions were found to be statistically significant. It was found that different genotype models increased Hb F levels between 1.6- and 3.06-fold in four studied polymorphic regions [rs4671393 (G>A), rs766432 (A>C), rs9402686 (G>A), rs28384513 (T>G)]. All of the polymorphic regions increased the Hb F levels from 1.86- to 24.76-fold, except rs9402686 (G>A) and rs28384513 (T>G) over dominant and rs1609812 (A>G) codominant inheritance models. The AC and AA genotypes increased Hb F levels in the B-cell CLL/lymphoma 11 A haplotype studies. It was determined that both haplotypes 2 and 4 increased Hb F levels. As a result, SNPs strongly affect the Hb F levels in both healthy individuals and β-thal trait.

Directed evolution of adenine base editors with increased activity and therapeutic application.

The foundational adenine base editors (for example, ABE7.10) enable programmable A•T to G•C point mutations but editing efficiencies can be low at challenging loci in primary human cells. Here we further evolve ABE7.10 using a library of adenosine deaminase variants to create ABE8s. At NGG protospacer adjacent motif (PAM) sites, ABE8s result in ~1.5× higher editing at protospacer positions A5-A7 and ~3.2× higher editing at positions A3-A4 and A8-A10 compared with ABE7.10. Non-NGG PAM variants have a ~4.2-fold overall higher on-target editing efficiency than ABE7.10. In human CD34+ cells, ABE8 can recreate a natural allele at the promoter of the γ-globin genes HBG1 and HBG2 with up to 60% efficiency, causing persistence of fetal hemoglobin. In primary human T cells, ABE8s achieve 98-99% target modification, which is maintained when multiplexed across three loci. Delivered as messenger RNA, ABE8s induce no significant levels of single guide RNA (sgRNA)-independent off-target adenine deamination in genomic DNA and very low levels of adenine deamination in cellular mRNA.

Molecular epidemiology and hematologic characterization of \u03b4\u03b2-thalassemia and hereditary persistence of fetal hemoglobin in 125,661 families of greater Guangzhou area, the metropolis of southern China

BackgroundIndividuals with δβ-thalassemia/HPFH and β-thalassemia usually present with intermedia or thalassemia major. No large-scale survey on HPFH/δβ-thalassemia in southern China has been reported to date. The purpose of this study was to examine the molecular epidemiology and hematologic characteristics of these disorders in Guangzhou, the largest city in Southern China, to offer advice for thalassemia screening programs and genetic counseling.MethodsA total of 125,661 couples participated in pregestational thalassemia screening. 654 subjects with fetal hemoglobin (HbF) level ≥ 5% were selected for further investigation. Gap-PCR combined with Multiplex ligation dependent probe amplification (MLPA) was used to screen for β-globin gene cluster deletions. Gene sequencing for the promoter region of HBG1 /HBG2 gene was performed for all those subjects.ResultsA total of 654 individuals had hemoglobin (HbF) levels≥5, and 0.12% of the couples were found to be heterozygous for HPFH/δβ-thalassemia, including Chinese Gγ (Aγδβ)0-thal, Southeast Asia HPFH (SEA-HPFH), Taiwanese deletion and Hb Lepore–Boston–Washington. The highest prevalence was observed in the Huadu district and the lowest in the Nansha district. Three cases were identified as carrying β-globin gene cluster deletions, which had not been previously reported. Two at-risk couples (0.0015%) were required to receive prenatal diagnosis. We also found 55cases of nondeletional-HPFH (nd-HPFH), including 54 with Italian nd-HPFH and one with the Aγ-197C-T heterozygous state. It is difficult to discriminate between Chinese Gγ (Aγδβ)0-thal and Italian nd-HPFH carriers using hemoglobin (Hb) analysis.ConclusionsThis study is the first to describe the familial prevalence of HPFH/δβ-thalassemia and the high-risk rate in Greater Guangzhou Area, and the findings will support the implementation of thalassemia screening for three common deletions by gap-PCR. We also presented a systematic description of genotype-phenotype relationships which will be useful for genetic counseling and prenatal diagnostic services for β-thalassemia intermedia.

Editing a γ-Globin Repressor Binding Site Restores Fetal Hemoglobin Synthesis and Corrects the Phenotype of Sickle Cell Disease Erythrocytes

Sickle cell disease (SCD) is a severe, life-threatening disorder caused by a single amino acid change (β6Glu→Val) in the adult hemoglobin (Hb) β-chain that causes Hb polymerization with consequent red blood cell (RBC) rigidity, anemia, vaso-occlusive crises, organ damage and reduced life expectancy. The co-inheritance of genetic mutations causing a sustained fetal γ-globin chain production in adult life (hereditary persistence of fetal hemoglobin, HPFH) decreases sickling and significantly reduces the clinical severity of SCD. HPFH mutations cluster at several loci in the promoters of the two γ-globin genes, HBG1 and HBG2, and, disrupt binding sites for transcriptional repressors (e.g., BCL11A and LRF), leading to elevated levels of fetal hemoglobin (HbF) representing up to 40% of total hemoglobin tetramers. In addition, SNPs at position -158 of both γ-globin promoters are associated with enhanced γ-globin expression and may identify a putative transcriptional repressor binding site. Here, we used the CRISPR/Cas9 system to mimic HPFH mutations in the HBG promoters by generating insertion and deletions (InDels) leading to disruption of known and putative binding sites for transcriptional repressors via non-homologous end joining (NHEJ) and microhomology-mediated end joining (MMEJ) DNA repair mechanisms. Efficient editing of the LRF binding site (≥ 3 γ-globin promoters/cell in &gt;70% of cells) in SCD patient-derived hematopoietic stem/progenitor cells (HSPCs) resulted in a robust, virtually pancellular HbF reactivation and a concomitant reduction in βS-globin levels, recapitulating the phenotype of asymptomatic SCD-HPFH patients. Of note, in RBCs derived from edited HSPCs, HbF levels exceeded 40% of total Hb tetramers, suggesting that CRISPR/Cas9 mediated disruption of the LRF binding site is even more potent than naturally occurring HPFH point mutations in reactivating HbF expression. RBCs derived from edited HSPCs displayed HbF levels sufficient to substantially correct the SCD cell phenotype (~80% of non-sickling RBCs). Although larger deletions (in part generated likely by MMEJ) disrupt more efficiently the LRF binding site, shorter InDels mainly produced by NHEJ, the most active repair mechanism in bona fide hematopoietic stem cells (HSCs), were still able to induce a potent γ-globin reactivation and correct the SCD cell phenotype. Interestingly, upon targeting of the -158 site, we observed HBG de-repression, which, however, was mild, indicating that this region contains a sequence that moderately inhibits γ-globin expression in adult cells, consistently with the mild increase in HbF associated with the -158 SNP. To evaluate the efficiency and the safety of this approach in long-term repopulating HSCs, untreated and edited HSPCs were injected into NSG immunodeficient mice. 16 weeks after transplantation, the engraftment of control and HBG-edited cells was comparable, as analyzed in bone marrow, spleen, thymus and blood. Edited HSCs were capable to differentiate into multiple hematopoietic cell types, with no skewing towards any particular lineage. Importantly, xenotransplantation of HSPCs edited using the gRNAs targeting the LRF binding site demonstrated a high editing efficiency in long-term repopulating HSCs (up to 75%) and a modest reduction in the frequency of deletions likely generated by MMEJ. Overall, this study identifies the binding site for the LRF repressor as a novel, potent and safe target for a genome editing-based treatment of SCD. Disclosures El Nemer: Hemanext: Other: Other. Cavazzana:SmartImmune: Other: Founder.

Genetic Modifiers of Sickle Cell Anemia in Patients from Nigeria: A Pilot Study

The variable clinical phenotype in sickle cell anemia is well documented, with severity in presentation partly associated with the persistence of fetal hemoglobin (HbF) and coinheritance of α-thala...

A Case of Hereditary Spherocytosis Caused by a Novel Homozygous Mutation in the SPTB Gene Misdiagnosed as β-Thalassemia Intermedia Due to a KLF1 Gene Mutation

We report a rare case of hereditary spherocytosis (HS) and hereditary persistence of fetal hemoglobin (Hb) (HPFH) complicated with a β-thalassemia (β-thal) trait and a Krüppel-like factor 1 (KLF1) gene mutation misdiagnosed as β-thal intermedia (β-TI) due to a high percentage of Hb F. The proband presented with pale skin, jaundice and splenomegaly. Analysis of the thalassemia gene indicated βcodon 17/βA (HBB: c.52A>T), while Hb analysis showed significantly increased Hb F levels. The proband was diagnosed to carry β-TI, and a blood transfusion regimen together with iron chelation treatment was recommended. Due to the difference between the phenotype and genotype, next generation sequencing (NGS) was performed and the proband was found to carry a homozygous mutation on the SPTB gene combined with a heterozygous mutation in KLF1. An eosin-5-maleimide binding test (EMA-BT) showed that the mean fluorescence intensity decreased by 47.1%. The proband was finally diagnosed with HS and HPFH complicated with a β-thal trait and the high percentage of Hb F was believed to be ascribed to the KLF1 gene mutation, which is frequent in areas where thalassemia is prevalent. For patients with a β gene mutation accompanying significantly high percentage of Hb F, the diagnosis of β-TI could be warranted, and the influence of the KLF1 gene mutation should be carefully excluded to avoid misdiagnosis of other types of hereditary hemolytic diseases.

Co-inheritance of Hereditary Persistence of Fetal Hemoglobin and Beta Thalassemia: A Rare Association

Co-inheritance of Hereditary Persistence of Fetal Hemoglobin and Beta Thalassemia: A Rare Association

A Rapid, Affordable and Feasible Method for Detection of the HBG1: g.-225_-222delAGCA Polymorphism

Single point mutations or small deletions in the Aγ - and Gγ-globin gene promoter region are associated to the nondeletional hereditary persistence of fetal hemoglobin (HPFH). Currently, DNA sequencing is most common technique adopted for detection of hemoglobin (Hb) mutations. However, some can be rapidly detected because they either destroy or create a recognition site for a restriction enzyme. Here we show that the 4 bp deletion, HBG1: g.-225_-222delAGCA in the Aγ-globin gene promoter can be easily detected using the Tru1I (MseI) restriction enzyme that cuts only in the absence of this deletion. This approach utilizes ordinary instrumentations (thermocycler and agarose gel electrophoresis) available in any basic molecular genetics laboratory, providing a reliable and inexpensive method of genetic screening.

The relationship between the variations in Gγ and Aγ promotors and the Hereditary Persistence of Fetal Hemoglobin (HPFH).

In the present study, sixty-two samples that have 1.5% and upper level of fetal hemoglobin (HbF), were examined to investigate the relationship between HbF level and non-deletional mutations in both Gγ (G gamma) globin (HBG2) and Aγ (A gamma) globin (HBG1) genes. Four variations were observed in the promotor of Gγ gene, which are -158C/T, -309A/G, -369C/G, and -567T/G. Also, four variations were observed in the 5'-UTR (untranslated regions) and promotor of Aγ gene, which are +25G/A, -369G/C, -499T/A, and -588G/A. One -222/-225 AGCA del homozygous and six variations as heterozygous in A gamma globin gene promotor region were also observed. The results of the current study suggested that there was a significant relationship between high HbF levels and two variations (-309A/T and -369C/G) in Gγ gene promotor. Additionally, a significant relationship between two variations (+25G/A and -499T/A) in Aγ gene promotor was also observed. Furthermore, the persons who carry these variations with high levels of HbF indicated that there might be a haplotype effect between these variations.

Spectrum of Types of Thalassemias and Hemoglobinopathies: Study in a Tertiary Level Children Hospital in Bangladesh

Thalassemias and hemoglobinopathies are the most common hemolytic congenital disorders in Bangladesh as in many parts of the world. This study was done to find the common types of thalassemias and abnormal hemoglobin variants seen in Bangladeshi populations. A total of 4813 samples were analyzed for hemoglobin disorders out of which 2308 (49.95%) showed abnormalities. The samples were analyzed by Bio Rad D 10 Analyzer in 3914 (81.32%) cases, BIORAD VARIANTβ thalassemia short program using the principle of high performance liquid chromatography in 474 (9.85%) cases and by CAPILLARYS 2 FLEX PIERCING utilizing capillary electrophoresis in 425 (8.83%) cases. The samples were analyzed in the Department of Biochemistry and Molecular Biology of Dhaka Shishu (Children) Hospital, Dhaka, Bangladesh. The common hemoglobin disorders seen were β trait 863 (17.94%), Hb E trait 601 (12.50%), Hb E β thalassemia 524 (10.87%), β thalassemia major 192 (4.00 %), Hb E disease 99 (2.05%). Other Hb abnormalities detected were Hb D trait 17 (0.35%), Sickle cell trait 4 (0.08%), hereditary persistence of fetal hemoglobin (HPFH) 2 (0.04%), and Hb Lepore, δ β thalassemia, sickle cell β thalassemia, Sickle cell disease, compound heterozygote for HbE+D and Hb Q band one case each (0.02%).

Haemoglobin F, A2, and S levels in subjects with or without sickle cell trait in south-eastern Gabon

ABSTRACTBackground: Infant mortality due to sickle cell disease in sub-Saharan Africa is high, necessitating a better understanding of the modulating factors of the disease in this region.Methods: We assessed the hereditary persistence of foetal haemoglobin and α-thalassemia. We diagnosed 787 subjects, with or without sickle cell trait, by capillary electrophoresis in the Medical Diagnostic Laboratory of the CIRMF (Franceville, Gabon).Results: Heterocellular and pancellular forms of hereditary persistence of foetal haemoglobin occurred at low rates of 10.9 and 2.3%, respectively. The distribution of HbS levels in individuals with sickle cell trait was trimodal, showing a high percentage (52.4%) of heterozygous subjects with α-thalassemia. The distribution of HbA2 levels was bimodal in individuals without sickle cell trait, estimated to be comprised of 12 and 15% of α and β-thalassemic heterozygous subjects, respectively.Conclusions: In sub-Saharan Africa, α-thalassemia is a far more prevalent modulating factor than hereditary persistence of foetal haemoglobin. Our study highlights the need for further investigation of thalassemia, haemoglobinopathies that are neglected in sub-Saharan Africa.