Cases Of Α-thalassemia Research Articles

Analysis of the Gene Mutation Type and Frequency of Thalassemia Patients in Jingzhou Area

To analyze the gene mutation types and frequence of thalassemia patients in Jingzhou area. A total of 721 suspected thalassemia patients who were visited in Jingzhou Central Hospital from June 2019 to June 2022 were selected as the research objects. There were 204 males and 517 females. PCR-reverse dot hybridization method was used to analyze the types and frequencies of 23 common α or β thalassemia gene mutations. Among the 721 patients with suspected thalassemia, 228 cases were positive for α or β thalassemia gene, with a total positive rate of 31.62%, including 87 cases of α-thalassemia, accounting for 38.16%, and 140 cases of β-thalassemia, accounting for 61.40%. There was 1 case of α β complex thalassemia, accounting for 0.44%. A total of 4 types of α-thalassemia gene mutations were detected, all of which were deletion types, including αα/--SEA (64/87, 73.56%), αα/-α3.7 (14/87, 16.09%), --SEA /-α3.7 (7/87, 8.05%), αα/-α4.2 (2/87, 2.30%). Among 140 patients with β-thalassemia, 138 were pure heterozygotes, and the genotypes of IVS-II-654M (63/140, 45.00%), CD41-42M (34/140, 24.29%), CD17M (18/140, 12.86%) and CD27-28M (10/140, 7.14%) accounted for 89.29% of all mutations (125/140), 2 cases of double heterozygosity (2/140, 1.43%) were found, no homozygous β-thalassemia were detected; 1 case of αβ complex thalassemia with genotype -α3.7/IVS-II-654M was found. The incidence of difference types of thalassemia was statistically significant (χ2=194.250, P < 0.001). The percentage of positive thalassemia genes was not significantly difference between male and female suspected patients (χ2=0.199, P =0.655). The α-thalassemia gene mutation in Jingzhou area is dominated by αα/--SEA, and the IVS-II-654M mutation is more common in β-thalassemia, and α β complex thalassemia is relatively rare, which can provide a reference for the formulation of prevention and treatment measures for thalassemia in Jingzhou area.

A particular focus on the prevalence of α-thalassemia and β-thalassemia among pregnant women in Changsha County, Hunan Province.

Thalassemia is a inherited monogenic blood disorder and more prevalent in southern China. In this study, Our aim was to elucidate the molecular spectrum and phenotypic features of thalassemia in pregnant women in Changsha County. Next-generation sequencing (NGS) was conducted for 38,810 pregnant women to diagnose thalassemia in Changsha County. Further analysis of hematological parameters was conducted on subjects who had not previously undergone thalassemia screening in other hospitals. In this study, 2,208 (5.69%) pregnant women were diagnosed as carriers of thalassemia using NGS analysis. Among 1,594 cases of α-thalassemia, 23 genotypes were identified, and among 578 cases of β-thalassemia, 22 genotypes were detected. Additionally, 18 genotypes were detected among 36 cases of composite α- and β-thalassemia. Among all carriers of thalassemia, 8 rare α-mutations and 11 rare β-mutations were found in the study population. Notably, pregnant women diagnosed as carriers of thalassemia tended to have lower hemoglobin levels. Furthermore, multivariable logistic regression analysis indicates that the values of MCV and MCH have the greatest impact on genetic diagnosis. Our study has provided detailed genotypes and hematological parameters of thalassemia in pregnant women in Changsha county and reveal that certain abnormal blood parameters have a remarkably impact on genetic diagnosis results. Furthermore, our data suggest that combining hemoglobin electrophoresis and NGS provides a powerful tool for prenatal diagnosis, which will increase the accuracy of clinical diagnosis of thalassemia.

Α-Globin Lentiviral Vectors for Hematopoietic Stem Cell Gene Therapy of α-Thalassemia

α-thalassemia is an inherited blood disorder caused by mutations in α-globin genes (HBA1 and HBA2) resulting in the reduction of α-globin chains, the subunit, along with β-globin chain, constituting adult hemoglobin (α 2β 2). While a reduction of α-globin expression of 40% presents asymptomatic phenotypes, expression levels decreasing to &amp;lt;20% or an α/β-globin mRNA ratio of &amp;lt;0.2, causes severe anima. Treatment for severe cases of α-thalassemia entails lifelong, biweekly blood transfusions with daily chelation therapy. While these therapies enable patients to live into mid- to late-adulthood, they continue to engender serious clinical manifestations, such as iron toxicity. To answer this clinical need, our laboratory has developed a stem cell gene therapy in which functional copies of α-globin gene integrate into the genome of patient's hematopoietic and progenitor stem cells (HSPCs) by lentiviral vectors (LVs) to normalize the globin chain imbalance and restore hemoglobin function in derived red blood cells (RBCs). The design of our α-globin LVs (AGLVs) is based on GLOBE β-globin LV utilized in a clinical trial for β-thalassemia (NCT02453477), which has achieved transfusion reduction and independence in patients with transfusion-dependence β-thalassemia major. We have constructed a series of short proviral length AGLVs for optimized titer production, HSPC infectivity, and lineage-specific α-globin gene expression. Twelve AGLVs varying in gene and regulatory element compositions were constructed and assessed for raw titer yields and characterized for gene transfer efficiency, mRNA expression and hemoglobin production in a α-globin knockout (KO) human erythroid cell line. Successfully, all AGLVs confer high raw titers ~ 1e7 TU/mL and enabled the production of adult hemoglobin (HbA). We identified two optimal AGLVs for further characterization in human primary HSPCs: 1) A2-UV for yielding highest raw titers and gene transfer efficiency, and 2) A2-Globe for producing the most α-globin mRNA. Both A2-UV and A2-Globe LVsharbor HBA2, are regulated by the β-globin promoter and enhanced by the core or large β-LCR enhancer region, respectively. To assess candidate AGLVs in human HSPCs, AGLVs were tagged at the transcription level to enable identification and quantification of vector-derived α-globin mRNA proceeding HSPCs transduction anderythroid differentiation. A2-UV demonstrated optimum CD34 + infectivity and gene transfer, and A2-Globe expressed a high ~30% of α-globin mRNA per total β-globin per vector copy number (VCN), achieving levels of one endogenous α-globin gene (~25% per total β-globin). Current studies examined A2-UV and A2-Globe in α-thalassemia major patient HSPCs - lacking all four α-globin genes - harboring a lethal homozygous 20kb deletion of HBA1 and HBA2, known as SEA –/– mutation. Patient HSPCs were transduced and differentiated into RBCs. A high CD34+ infectivity and gene transfer was obtained by both candidate vectors, without utilization of transduction enhancers. Furthermore, a correction of α/β-globin mRNA ratio of ~0.7 was obtained while retaining a low VCN (&amp;lt;3).RBCs were then analyzed at the protein level . Chromatograms of hemoglobin tetramers revealed that HbA represented 98% of all hemoglobin, compared to 0% in non-transduced patient RBCs. Single globin chain chromatograms successfully demonstrated α/β-globin chain ratio of ~0.4 or higher and successfully demonstrated that a low average of copies per cell (~3) restored 60% of adult hemoglobin when normalized to healthy donor RBCs. These data demonstrate α/β-globin correction and hemoglobin restoration in α-thalassemia patient cells with the most severe mutation. Three assays were conducted on patient cells, all with similar and highly encouraging results for disease amelioration in severe cases of α-thalassemia. Overall, this project is paving the way for the first development of a curative gene therapy approach for α-thalassemia.

Molecular Spectrum, Ethnic and Geographical Distribution of Thalassemia in the Southern Area of Hainan, China.

BackgroundThalassemia is one of the most common genetic diseases in southern China. Accurate population frequency data regarding the occurrence and distribution of thalassemia are important for designing appropriate prevention strategies for thalassemia. This study aims to reveal the molecular spectrum, ethnic and geographical distribution of thalassemia in the southern area of Hainan Province, China.MethodsA total of 9813 suspected carriers of thalassemia were screened for genetic analysis by using the PCR-reverse dot blot hybridization method targeting three known deletions of α-thalassemias (--SEA, -α3.7, and -α4.2), three nondeletional mutations of α-thalassaemias (αCS, αQS, and αWS) and the 17 most common mutations of β-thalassaemias in the Chinese population.ResultsApproximately 6,924 subjects were genetically diagnosed as thalassemia carriers or patients, including 5812 cases of α-thalassemia (83.9%), 369 cases of β-thalassemia (5.3%), and 743 cases of α-composite β-thalassemia (10.7%). A total of 21 distinct genotypes were identified among the 5,812 α-thalassemia carriers, -α4.2/αα, -α3.7/αα, and -α3.7/-α4.2 were the most common α-thalassemia genotypes. The most frequent β-thalassemia genotype was βCD41−42/βN, with a notable proportion of 69.6%, followed by the β−28M/βN, βIVS−II−654/βN, βCD71−72/βN, βE/βN, and βCD17/βN genotypes. In addition, 37 genotypes were detected among the 743 cases of both α- and β-thalassemia mutations. The α-thalassemia genotypes were most commonly found in the Li people, who accounted for 73.5% of α-thalassemia carriers. The β-thalassemia genotypes were most commonly identified in the Han people, who accounted for 59.4% of β-thalassemia carriers. Among the subjects carrying both α- and β-thalassemia variations, only three ethnic minorities were identified, including the Li, Han, and Miao people, accounting for 82.0, 17.4, and 0.7%, respectively.ConclusionsOur study indicates that there is high genetic heterogeneity, geographical and ethnic differences in thalassemia in populations in the southern area of Hainan Province. These findings will be helpful in guiding genetic counseling and prenatal diagnosis of thalassemia in Hainan Province.

Molecular Spectrum of α- and β-Thalassemia among Young Individuals of Marriageable Age in Guangdong Province, China

Thalassemia is a group of genetically heterogeneous diseases characterized by hemolytic anemia. To investigate molecular characteristics of α- and β-thalassemia among young individuals of marriageable age in Guangdong Province, 24,788 subjects with suspected thalassemia were genetically tested for α- and β-thalassemia by Gap-PCR and reverse dot blot during 2018-2019. For suspected rare thalassemia cases, DNA sequencing was performed to identify rare and unknown thalassemia gene mutations. A total of 14,346 thalassemia carriers were detected, including 7,556 cases of α-thalassemia with 25 genotypes and 8 α-gene mutations identified, 5,860 cases of β-thalassemia with 18 genotypes and 18 β-gene mutations identified, and 930 cases of compound α/β-thalassemia. Among them, the frequency of -- SEA deletion was the highest in α-thalassemia (66.01%), followed by -α 3.7 (17.98%) and -α 4.2 (8.22%), and the frequency of CD41-42 (-TCTT) mutation was the highest in β-thalassemia (38.38%), followed by IVS-II-654 (C > T) (25.67%), -28 (A > G) (15.76%), and CD17 (10.01%). In addition, 5 rare mutations (--THAI and HKαα, CD113, -90, and CD56) were found in the study population. Our results revealed molecular epidemiological background of α- and β-thalassemia in Guangdong Province, which can support optimization of thalassemia prevention and control strategies. We demonstrated that thalassemia is heterogeneous with significant geographical differences and population specificity.

Gene Detection Analysis of Thalassemia in 2 494 Cases of Childbearing Couples in Haikou City

To observe the genotypes and composition ratio of thalassemia in couples of reproductive age, and provide a reference for the prevention and control of thalassemia in Haikou. Gene diagnosis was performed in 2 494 subjects who were screened for thalassemia before marriage or prenatal by cross-breakpoint PCR, PCR-reverse dot hybridization, and PCR-electrophoresis. A total of 1 037 thalassemia gene carriers were detected in 2 494 samples, with a detection rate of 41.57%, of which 75.02% was α-thalassemia, 18.61% was β-thalassemia, and 6.36% was α-β complex thalassemia. There were 778 cases of α-thalassemia, mainly of deletion type, accounting for 76.99% (599/778). Twenty genotypes were detected, the highest three was --SEA/αα (33.42%, 260/778), -α3.7/αα(23.91%, 186/778), and -α4.2/αα(19.02%, 148/778), respectively. A rare HKαα/-α3.7 was detected, who immigrated from other province. There were 193 cases of β-thalassemia, all of them were light (β0/βA or β+/βA). Eight genotypes were detected, the highest two was 41-42M/N (74.61%, 144/193) and 654M/N (10.36%, 20/193), respectively. There were 66 cases of α-β compound type of thalassemia, 15 genotypes were detected, the highest three was ααWS/αα complex 41-42M/N (28.79%, 19/66), -α3.7/αα complex 41-42M/N, and -α4.2/αα complex 41-42M/N (16.67%, 11/66 for both). In Haikou city, the gene carrying rate of thalassemia is very high, and the genotype distribution is different from other cities in Hainan Province, attention should be paid to the impact of population inflow on the frequency spectrum change of local thalassemia gene.

Analysis of Gene Mutation Types in 141 Cases of α-Thalassemia in Honghe Prefecture, Yunnan Province

To detecte the carrying rate, the type and distribution of α-Thalassemia gene mutation in Honghe Prefecture, Yunnan Province, and analyze the differences in average erythrocyte volume (MCV), mean erythrocyte hemoglobin content (MCH) and hemoglobin among different types of α-Thalassemia. The DNA samples from small cell hypochromic carriers or anemia patients and women of childbearing age who underwent hematological screening in The First People's Hospital of Honghe State was from 2015 to 2019 were enrolled and analyzed, and the mutation types and frequency of alpha-thalassemia positive rate were diagnosed by PCR reverse dot blot or PCR fluorescence dissolution curve. Among the 1 016 samples, 141(13.88%) of the patients were diagnosed as α-thalassemia. The α-thalassemia was subdivided into 3 types, silent (36.17%), minor (51.77%), and HbH disease (12.06%), and the MCV, MCH and HB levels were detected and showed a obvious decrease trend with significant difference (P < 0.05). The gene mutation types were 9 kinds, the deletion type gene was mainly --SEA (51.06%), followed by -α3.7/αα deletion (29.79%), the α- mutation type gene was mainly αcsα(3.55%). The absence of complex heterozygote was most common, which was 17 cases, accounting for 12.77%, mainly --SEA/-α3.7 (10.64%). The areas were mainly distributed in Mengzi (47.52%), followed by Jinping (17.02%). Ethnic groups were mainly distributed in Han nationality (49.65%), followed by Zhuang (15.60%), Yi (3.48%) and Dai (7.09%). The patients with double heterozygous mutation was slightly higher in Mengzi than that in Jinping County (4.26% vs 3.55%), the Dai (2.13%) showed Zhuang ethnic group (2.13%) had the relatively high incidence except the Hans, and showed the most serious anemia. Alpha-thalassemia in Honghe prefecture of Yunnan Province shows complex genetic diversity and significant genetic heterogeneity, and the mainly type of gene mutation is --SEA and --SEA/-α3.7, which is mainly distributed in Han, Zhuang and Dai ethnic groups in Mengzi, Jinping. The anemia index of HbH group is the most obvious, and it is significantly different from other groups.

Early genetic screening uncovered a high prevalence of thalassemia among 18 309 neonates in Guizhou, China

Thalassemia is a common monogenic disease in southwestern China, especially in Guizhou province. In this study, 18 309 neonates were examined for thalassemia. The thalassemia carrier rate was 12.90%, which is associated with geographical regions, with carrier frequencies significantly differing between regions (p < 0.0001). The carrier rates for α-thalassemia and β-thalassemia were 8.91% and 3.36%, respectively. There are 22 genotypes identified among 1632 α-thalassemia cases, and 18 genotypes detected among 615 β-thalassemia cases. The birthrates of individuals with intermediate thalassemia and β-thalassemia major were 0.153% and 0.055%, respectively. Methodologically, NGS-Gap-PCR is superior to traditional detection methods, with 65 more cases detected by NGS-Gap-PCR. Since thalassemia-rich genotypes were highly prevalent in this region, early detection of thalassemia carriers would be meaningful for genetic counseling and prevention/treatment of thalassemia. NGS-Gap-PCR provides a powerful tool for neonate genetic testing and clinical diagnosis of thalassemia, especially in high-prevalence regions.

Clinical value of PCR-flow Fluorescence Hybridization in Prenatal Genetic Diagnosis of Thalassemia

To explore the value of PCR-flow fluorenscence immunmicrobeads assay in prenatal gene diagnosis of thalassemia. A total of 1001 pregnant women and their couples checked in the First Affiliated Hospital of Sun Yat-Sen University from January 2016 to August 2019 were selected. Both pregnant women and their spouses were the carriers of thalassemia gene. Samples such as amniotic fluid, were used to extract genomic DNA at the right time. Parallel detection of α- and β- thalassemia genes to samples should be carried out by PCR-flow cytometric fluorescence hybridization and traditional multiple Gap-PCR and PCR-RDB techniques. The consistency of two methods in gene diagnosis of thalassemia was evaluated by analyzing the results of detection. 389 normal genotypes (38.86%, 389/1001) and 59 abnormal genotypes (61.14%, 612/1001) was cheked out by the two methods, including 416 cases of α-thalassemia, 162 cases of β-thalassemia and 34 cases of αβ- complex thalassemia. The main genotypes of α-thalassemia were --SEA, -α3.7 and -α4.2. The mutation frequency of CD41-42 was the highest among the β-thalassemia genotypes, which followed by IVS-II-654 and CD17. A rare HKαα/--SEA thalassemia genotype was detected. Compared the traditional multiple Gap-PCR and PCR-RDB techniques, the sensitivity, specificity, positive predictive value, negative predictive value and total consistent rate of PCR-flow fluorenscence immunmicrobeads assay were 100%, which showed that the two methods were completely consistent. Guangzhou is a area with high incidence of thalassemia, and the genetic types of thalassemia are complex and diverse. Prenatal diagnosis is the final barrier to the prevention of thalassemia. PCR flow-cytometric fluorescence hybridization, as a simple and fast technique, combined with traditional techniques in parallel contributed to the accuracy of prenatal gene diagnosis of thalassemia.

Gene characteristics and changing trend of neonatal thalassemia in Dongguan, China

To study the gene distribution characteristics of neonatal thalassemia in Dongguan, China and the changing trend of the gene distribution characteristics of neonates with thalassemia in Dongguan in 2014-2018. A retrospective analysis was performed for the data on neonatal thalassemia screening from the Dongguan Neonatal Disease Screening System between January 2014 and December 2018. A total of 616 718 neonates were enrolled who were born in Dongguan. Among the 616 718 neonates, 52 308 were positive for primary screening, 10 366 were recalled, 8 576 underwent genetic diagnosis, and 6 432 were confirmed with thalassemia by genetic diagnosis. The carrying rates of thalassemia genes in 2014-2018 were 5.81%, 5.47%, 5.96%, 6.91%, and 7.90% respectively, and showed an upward trend (P<0.001). The positive rates of neonatal thalassemia screening in 2014-2018 were 9.12%, 8.34%, 7.54%, 8.13%, and 9.32% respectively (P<0.001). The positive rates of genetic diagnosis of neonatal thalassemia in 2014-2018 were 0.89%, 1.11%, 1.24%, 0.90%, and 1.09% respectively (P<0.001). In 2014-2018, 5 098 cases of α-thalassemia were detected, accounting for 79.26% of all cases, and 1 230 cases of β-thalassemia were detected, accounting for 19.12% of all cases. The detection rate of α-thalassemia was significantly higher than that of β-thalassemia in each year (P<0.001). In 2014-2018, static α-thalassemia, mild α-thalassemia, and mild β-thalassemia were the main types observed in neonates. Most of the neonates with thalassemia have α-thalassemia in Dongguan, with static α-thalassemia and mild α-thalassemia as the main types. The carrying rate of thalassemia genes keeps increasing in neonates in Dongguan, and the prevention and treatment of thalassemia is still challenging.

Analysis on the Genotype of 5018 Cases of Thalassemia in Hunan Area

To investigate the type and distritution of thalassemia gene mutaitons in Hunan area, so as to provide evidence for prenatal screening, diagnosis and reduction of birth defects. A total of 5018 cases from Maternal and Children Health Hospital of Hunan from June 2017 to Dec 2018 were undergone thalassemia gene mutation analysis. The reverse dot blot hydridization was used to detect 6 kinds of genotypes of α-thalassemia and 17 kinds of point mutations of β-thalassemia, and the detected data were analyzed statistically. 889 cases (55.9%) of α-thalassemia carriers were found, including 385 cases of silent α-thalassemia, 488 cases of α-thalassemia trait, 16 cases of Hb H disease. --SEA/αα was the most common genotype in α thalassemia. 664 cases (41.7%) were diagnosed as β-thalassemia carriers, heterozygotes accounted for 99.8% (663/664), IVS-Ⅱ-654, CD41-42M and CD17M were the main genotypes, and compound heterozygote accounted for 0.2% (1/664). 38 cases were diagnosed as α-thalassemia combined β-thalassemia. The constituent ratio of thalassemia gene mutations in Hunan has regional characteristics, --SEA/αα is the most common genotype in α-thalassemia carrier. IVS-Ⅱ-654, CD41-42 and CD17 are common ones in β-thalassemia. The frequency of α-thalassemia combined with β-thalassemia is high.

Analysis on the Genotype of 5018 Cases of Thalassemia in Hunan Area

To investigate the type and distritution of thalassemia gene mutaitons in Hunan area, so as to provide evidence for prenatal screening, diagnosis and reduction of birth defects. A total of 5018 cases from Maternal and Children Health Hospital of Hunan from June 2017 to Dec 2018 were undergone thalassemia gene mutation analysis. The reverse dot blot hydridization was used to detect 6 kinds of genotypes of α-thalassemia and 17 kinds of point mutations of β-thalassemia, and the detected data were analyzed statistically. 889 cases (55.9%) of α-thalassemia carriers were found, including 385 cases of silent α-thalassemia, 488 cases of α-thalassemia trait, 16 cases of Hb H disease. --SEA/αα was the most common genotype in α thalassemia. 664 cases (41.7%) were diagnosed as β-thalassemia carriers, heterozygotes accounted for 99.8% (663/664), IVS-Ⅱ-654, CD41-42M and CD17M were the main genotypes, and compound heterozygote accounted for 0.2% (1/664). 38 cases were diagnosed as α-thalassemia combined β-thalassemia. The constituent ratio of thalassemia gene mutations in Hunan has regional characteristics, --SEA/αα is the most common genotype in α-thalassemia carrier. IVS-Ⅱ-654, CD41-42 and CD17 are common ones in β-thalassemia. The frequency of α-thalassemia combined with β-thalassemia is high.

Impact of the detection of ζ-globin chains and hemoglobin Bart's using immunochromatographic strip tests for α0-thalassemia (--SEA) differential diagnosis.

α0-Thalassemia is an inherited hematological disorder caused by the deletion of α-globin genes. The Southeast Asian deletion (--SEA) is the most common type of α0-thalassemia observed in Southeast Asian countries. Regarding WHO health policy, an effective α0-thalassemia screening strategy is needed to control new severe α-thalassemia cases. In this study, a monoclonal antibody panel was used to develop immunochromatographic (IC) strip tests for detecting the Hb Bart’s and ζ-globin chain. Among 195 samples, all α0-thalassemia traits (78 α0-thalassemia (--SEA) and 4 α0-thalassemia (--THAI)) had low MCV or MCH values. The sensitivity, specificity, PPV and NPV of the IC strip tests for ζ-globin and Hb Bart’s for screening α0-thalassemia (--SEA) within the low MCV or MCH samples were 100%, 65.2%, 90.7%, 100% and 96.2%, 47.8%, 86.6%, 78.6%, respectively. All 4 α0-thalassemia (--THAI) traits were negative for ζ-globin chains but positive for Hb Bart’s using the IC strip tests. These results led to a α0-thalassemia screening being proposed in which blood samples are first evaluated by MCV, MCH and Hb typing. Samples with high MCV and MCH values are excluded for the presence of the α0-thalassemia gene. Samples with low MCV or MCH values are assayed using the developed IC strip tests, where only samples testing positive are further assayed for α0-thalassemia by PCR. Patients with Hb H, EA Bart’s or EF Bart’s diseases do not need to use this IC strip assay. Thus, in this study, a simple and cost effective α0-thalassemia point of care test was developed.

Prevalence and genetic analysis of thalassemia in neonates in Wuhan area: a national megacity in central China

Background Thalassemia is one of the most common genetic diseases in southern China. Accurate population frequency data regarding the occurrence and distribution of thalassemia is important for designing appropriate prevention strategies of thalassemia. Objective The aim of this study is to reveal the prevalence and the mutation spectrum of thalassemia in neonates in the Wuhan region of central China. Methods About 3796 neonates in Wuhan area of China were analyzed by hematological and genetic analysis. Results About 2174 subjects were genetically diagnosed as thalassemia carriers or patients, including 1415 cases of α-thalassemia (65.89%), 731 cases of β-thalassemia (33.62%), and 28 cases of α-composite β-thalassemia (1.29%). A total of 11 genotypes and 6 gene mutations were identified in α-thalassemia anomalies, with —SEA/deletion (50.72%), -α3 . 7/deletion (36.36%), and -α4 . 2/deletion (7.38%) being the most common α-thalassemia mutations. β-thalassemia anomalies were associated with 17 genotypes and 12 gene mutations; IVS-2-654 mutation was the most common (41.18%), followed by CD41-42 (23.14%), CD17 (14.64%), CD26 (7.32%), and CD27-28 (4.58%) mutations. In addition, 13 genotypes were identified in α-composite β-thalassemia in thalassemia carrier, with the top six genotypes being IVS-2-654/N/—SEA/αα (17.86%), CD17/N/-α3 . 7/αα (17.86%), IVS-2-654/N/-α3 . 7/αα (14.29%), CD41-42/N/—SEA/αα (10.71%), CD71-72/N/-α3 . 7/αα (7.14%), and Cap/N/—SEA/αα (7.14%). Conclusion There was high heterogeneity and extensive spectrum of thalassemia in the neonates in Wuhan populations. The findings will be useful for genetic counseling and prenatal diagnosis of thalassemia in the Wuhan region.

Cut-Off Values of Hematologic Parameters to Predict the Number of Alpha Genes Deleted in Subjects with Deletional Alpha Thalassemia.

Most α-thalassemia cases are caused by deletions of the structural α-globin genes. The degree of microcytosis and hypochromia has been correlated with the number of affected α-globin genes, suggesting a promising role of hematologic parameters as predictive diagnostic tools. However, cut-off points for these parameters to discriminate between the different subtypes of α-thalassemia are yet to be clearly defined. Six hematologic parameters (RBC, Hb, MCV, MCH, MCHC and RDW) were evaluated in 129 cases of deletional α-thalassemia (56 heterozygous α+ thalassemia, 36 homozygous α+ thalassemia, 29 heterozygous α0 thalassemia and 8 cases of Hb H disease). A good correlation between the number of deleted alpha genes and MCV (r = −0.672, p < 0.001), MCH (r = −0.788, p < 0.001) and RDW (r = 0.633, p < 0.001) was observed. The presence of an α0 allele should be discarded in individuals with microcytosis without iron deficiency and normal values of Hb A2 and Hb F with MCH < 23.40 pg. Furthermore, MCH < 21.90 pg and/or MCV < 70.80 fL are strongly suggestive of the presence of one α0 allele. Finally, an accurate presumptive diagnosis of Hb H disease can be made if both RDW ≥ 20% and MCH < 19 pg are seen.

α-globin Alteration in α-thalassemia Disorder: Prediction and Interaction Defect.

The α-thalassemia is an inherited blood disorder affecting quality and quantity of hemoglobin. It caused mostly by deletion of one or two α-globin genes and characterized by deficient production of α-globin chain in hemoglobin leading from mild anemia to lethal. The α-globin gene with partial deletion could reduce chain production or produce abnormal chain. Its effect depends on mechanism of chain production affected. This study aimed to analyze the effect of partial deletion in α-globin gene influencing the mechanisms to produce functional α-globin chain in α-thalassemia cases. The three mutant genes from genebank were selected and processed. The analysis performed in deleted sequences determination, mRNA sequences, protein structures and protein chains interaction to form hemoglobin by SWISS MODEL, CHIMERA and SABLE Polyview 2D. The result showed 76 amino acids deleted in one mutant α-globin gene (V00516.1). The mutation gave effect in every mechanism of the α-globin chain conformation and production. It affected protein conformation by losing over half the helical chains. It reduced the function completely, in which, disturb hemoglobin A (HbA) production with emergence of β-sheets conformation. The analysis concluded that the protein produced by the α-globin gene with partial deletion lost its function and unable to form hemoglobin.

Molecular mechanism of yisui shengxue granule, a complex chinese medicine, on thalassemia patients suffering from hemolysis and anemia of erythrocytes.

The objective of this study was to investigate the therapeutic biological mechanism of Yisui Shengxue Granule (YSSXG), a complex Chinese medicine, on the hemolysis and anemia of erythrocytes from patient with thalassemia disease. Sixteen patients with thalassemia (8 cases of α-thalassemia and 8 cases of β-thalassemia) disease were collected and treated with YSSXG for 3 months. The improvements of blood parameter demonstrated that YSSXG had a positive clinical effect on patients with thalassemia disease. For patients with α-thalassemia disease, RT-PCR showed that YSSXG upregulated the relative mRNA expression level of α-globin to β-globin and downregulated DNMT1, DNMT3a, and DNMT3b mRNA compared with pretreatment. Western blotting showed that YSSXG downregulated the expression of DNMT1 and DNMT3a. For patients with β-thalassemia disease, the relative expression level of A γ-globin to α-globin had an increasing trend and the level of BCL11A mRNA expression obviously increased. For all patients, RT-PCR showed that YSSXG upregulated mRNA expression of SPTA1 and SPTB. Activities of SOD and GSH-Px significantly increased and MDA obviously reduced on erythrocyte and blood serum after YSSXG treatment. TEM showed that YSSXG decreased the content of inclusion bodies. Activities of Na+K+-ATPtase and T-ATPtase of erythrocyte increased significantly after YSSXG treatment. This study provides the basis for mechanisms of YSSXG on thalassemia suffering with hemolysis and anemia of erythrocytes from patient.

The Contribution of Extramedullary Hematopoiesis to Hepatomegaly in Anemic Hydrops Fetalis: A Study in Alpha-Thalassemia Hydrops Fetalis

Massive hepatomegaly is a common finding in hydrops fetalis (HF) arising from hemoglobinopathies. It has been suggested that extramedullary hematopoiesis, which is markedly increased in response to anemia, plays a principal role in hepatomegaly via sinusoidal obstruction and distortion of the intrahepatic architecture. To test this concept, the authors compared 20 cases of α-thalassemia (hemoglobin Bart, 15 cases; hemoglobin H, 5 cases) with 19 cases of HF arising from other causes and 39 control nonhydrops cases. The mean liver weight in α-thalassemia cases was significantly heavier than in control cases and in hydrops fetalis cases due to other causes (P < 0.01). This was not explainable on the basis of extramedullary hematopoiesis (erythropoiesis or myelopoiesis) or hemosiderin deposition, since cases of HF from other causes, especially related to anemia, showed similar findings to α-thalassemia cases. While these processes no doubt contribute to hepatic weight, the major factor would still be high-output cardiac failure causing hepatic congestion.

Molecular characterization of a novel 27.6-kb deletion causing α+ thalassemia in a Chinese family

Over 80% of the α-thalassemia cases in southern China are caused by large deletions involving the α-globin gene cluster on chromosome 16p13.3. Here, we characterized a novel 27.6-kb deletion on the α-globin gene cluster in a Chinese family. Its breakpoints were detected to lie between coordinates 9079 and 36718 of the α-globin gene cluster (NG_000006.1), with a total of 27,640 nucleotides deleted. It was designated as -α (27.6) deletion. The proband is a compound heterozygote of --(SEA) and -α (27.6) and he displayed very mild hemoglobin H disease phenotype with Hb 7.9-9.3 g/dl. Phenotypic analysis on heterozygote of this deletion revealed it as α(+) mutation. It leads to a very mild phenotype as adult heterozygotes have normal hematological parameters with the values at the lower border of the normal range. RT-PCR analysis showed that the α-globin mRNA level of the heterozygotes was decreased when compared with that of normal people.

Relevance of α-Globin Genotyping in Patients Expressing HbH in Cord Blood: Prevalence of α-Globin Gene Deletions and α2-Globin Termination Codon Mutations in a Neonatal Hemoglobinopathy Screening Program

Introduction: α-Thalassemia results from an heterogeneous group of disorders involving the α-Globin genes. Common deletions are responsible for 95% of cases of α-thalassemia, whereas Constant Spring mutation (Hb CS), one of the known mutations involving the termination codon of α2-globin gene, is the most common non-deletional cause of α-thalassemia worldwide. Its prevalence has never been studied in occidental countries.Patients and methods: as part of our neonatal hemoglobinopathy screening program, fetal cord blood samples are systematically tested by HPLC. When an abnormal Hb or HbH is found, α-Globin deletion is carried by multiplex-PCR. A two part observational study was done at our center based on the available data from our hemoglobinopathy screening program available since 2002. Firstly, we aimed to determine the level of Hb H present and to correlate it to a specific type of α-Globin gene deletion by m-PCR sequencing (table 1). 306 samples were analyzed and demonstrated a statistically significant difference between categories of α-globin gene deletions: 2 del cis vs 1 del, 2 del trans vs 1 del, 2 del cis vs 2 del trans (difference of means of 8.07, 4.33 and 3.75 respectively, P<0.001). With HbH ≤3.15% a CI of 95% excludes a condition other that 1 α -globin gene deletion. With Hb H ≤7.1%, a CI of 99% excludes 2 α -globin gene deletions in cis.Table 1Deletion/allelic distributionHb H level (range)Hb H level (means)Std Dev1 deletion1.0 – 12.42.031.122 deletions cis-10.12.162 deletions trans-6.351.5Our second goal was to establish the prevalence of α-Globin termination codon mutation in the fetal blood sampling containing ≥1% of Hb H in absence of α-Globin gene deletion by m-PCR. 395 fetal blood samples were analysed. Of these, 59 lacked α-Globin gene deletions. 17 were excluded from our analysis for lack of personal data or DNA. The 42 remaining samples were studied by amplification of the termination codon followed by an enzymatic digestion for the recognition site TTAA. 2 samples (5%) were found to be heterozygotes for α2-globin mutation, presumably CS mutation. The small number of positive cases does not permit the conduct of a powerful statistical analysis, however the two mutated samples had a high level of Hb H (2,9%) whereas only 21% of the 42 studied samples had an Hb H ≥2%.Conclusion: These results represent a step forward in diagnosis of α-thalassemia and in identification of patients requiring particular genetic counselling based on HbH level at birth in cord blood. These results also indicate that 5% of Hb H positive/α-Globin deletion negative samples were caused by termination codon mutation in an occidental urban based population, and that it could be of interest to search for these mutations, especially when the Hb H level is high.