Blood Transfusion-dependent Patients Research Articles

Mutation Spectrum of β-Globin Gene in Patients with β-Thalassemia at Tidar Hospital, Magelang, Central Java, Indonesia

β-Thalassemia is genetic disorder characterized by β-globin chain deficiency resulting from mutations in the β-globin coding gene. Both the quantity and quality of blood produced will be impacted by this condition. The distribution of mutation causing thalassemia is vary across ethnic and different regions in Indonesia. This study aims to identify the variant mutation in patients with β-thalassemia at Tidar Hospital as representative samples of Javanese population, the largest ethnicity in Indonesia. Sixty-one blood samples were obtained from blood transfusion-dependent patients with β-thalassemia. Mutation was identified using ARMS and RFLP PCR-based methods, and inconclusive samples were subjected to DNA sequencing. Results showed that the mutation variants were Cd 26/IVSI-5 (G > C) 47.54%, Cd 26/Cd 35 16.30%, Cd 26/IVSI-1 (G > T) 11.47%, Cd 26/IVSI-2 4.91%, IVSI-5 (G > C)/Cd 40 3.27%; 1.63%; IVSI-5 (G > C)/IVSI-1 (G > A) 1.63%; IVSI-5 (G > C)/Cap + 1 1.63%; Cd 26/Cd 15 1.63%; Cd 26/Cd 30 1.63%. We also found three homozygous of IVSI-1 (G > T), IVSI-5 (G > C) 6.55%, and Cd 35 1.63%. The most prevalent alleles would be recommended to be used as part of screening for β-thalassemia in the Javanese ethnicity in Central Java, especially for families affected by thalassemia.

Assessment of orthodontic treatment needs in patients with β-thalassemia major and sickle cell disease.

Patients suffering from hemoglobinopathies may suffer from pathogic dental and orofacial features. This study aimed to assess the prevalence of malocclusion and the need for orthodontic treatment in patients with β-thalassemia major (BTM) and sickle cell disease (SCD). The study was conducted on 311 blood transfusion-dependent patients with BTM or SCD and 400 healthy individuals aged 10 to 16. The types of malocclusion were evaluated based on Angle's classification and Dewey's modification, and their oral habits were recorded using a questionnaire. The need for orthodontic treatment was assessed through the Dental Health Component of the Index of Orthodontic Treatment Need (IOTN), and the data were compared with normal participants. The Index of Orthodontic Treatment Need-Dental Health Component (IOTN-DHC) assessment showed that patients had a higher prevalence of objective need for treatment (IOTN grades 4 and 5) compared to healthy children. The prevalence of class II malocclusion was significantly higher in patients. Patients showed significantly less Angle's Class I malocclusion compared to normal participants. Oral habits were presented in 61%, 64.15% and 62.4% of normal participants, BTM and SCD patients, respectively. The higher prevalence of Angle's class II malocclusion and higher percentage of IOTN grade 4 and 5 among BTM and SCD patients reveal the importance of early orthodontic assessment and intervention in children with BMT and SDC.

The application value of ultra-short echo time MRI in the quantification of liver iron overload in a rat model

The quantitative evaluation of liver iron concentration (LIC) is important in guiding the treatment of blood transfusion-dependent patients. Conventionally, LIC is assessed through R2*or R2 values using magnetic resonance imaging (MRI). However, most of the studies using MRI to determine iron overload were restricted by the minimum echo time, so that severe iron overload could hardly be quantified. In our study, we demonstrate a new approach to overcome the limitation of the shortest echo time using ultra-short echo time (UTE) MRI to quantify liver iron overload of varying degrees in a rat model. Sixty female Sprague-Dawley rats were included and randomly assigned into 10 equal groups. Group 1 was not injected with iron dextran. Groups 2 to 10 were intraperitoneally injected with iron dextran at a dose of 15 mg/kg every 3 days. On every 6th day, one group was randomly selected from groups 2 to 10 for MRI scanning and liver iron concentration (LIC) detection. For groups 1 to 10, images were acquired by UTE sequence using a 3.0T MR scanner, and the T2* value and R2* value were obtained (R2* =1/T2*). In addition, LIC was measured using an atomic absorption photometer. The correlation analysis between R2* value and LIC was performed and the regression equation of R2* and LIC was established and its reliability verified. For groups 1 to 10, R2* values and LIC ranged from 60.16±4.76 to 1,306.90±42.26 Hz and from 0.84±0.11 to 5.89±2.64 mg/g dry, respectively. The R2* value was linearly correlated to the LIC (r=0.897, P<0.001), and the linear regression equation was LIC = 0.005 × R2* + 1.783. The validation analysis results showed that the intragroup correlation coefficient (ICC) between the predicted and measured LIC was 89.5%. The UTE sequence could be used for quantification of varying degrees of hepatic iron overload in the rat model, and the LIC could be predicted by using the R2* value on an MR 3.0T scanner.

Evaluation of Liver Iron Deposition in Transfusion-Dependent Patients By Dual-Energy CT

▪[Introduction]The launch of the oral iron chelator "Deferasirox" has improved the outcomes of blood transfusion-dependent patients with iron overload in the last decade. Although serum ferritin (SF) remains the mostly commonly used metric to monitor body iron stores for decisions regarding the indication of iron chelate therapy, it is known to be affected by many factors. The liver iron concentration (LIC) is considered to be an indicator of total body iron stores, and the MR imaging-based R2 technique is the standard non-invasive technique used to evaluate LIC. However, this technique is not used in every institution due to some limitations such as its high cost and the requirement for special software. Although the application of CT, which is easy to use and inexpensive, needs to be considered for the evaluation of LIC, the use of conventional single energy CT (SECT) to measure LIC is also limited by normal variations in CT attenuation, predominantly in patients with mild iron overload. Moreover, SECT fails to detect iron in fatty livers, which has an inverse effect on attenuation by lowering CT numbers. Dual-energy CT (DECT) is a technique that is employed to obtain precise information on tissue composition and may be useful for monitoring LIC. It is based on substances showing different densities with two different energies, with each substance displaying its own energy-dependent change in CT attenuation. The role of DECT in monitoring LIC has not yet been clarified in blood transfusion-dependent patients with iron overload. We herein evaluated iron deposition in the livers of blood transfusion-dependent patients using DECT.[Patients and Methods]Seventeen blood transfusion-dependent patients underwent liver DECT using a dual-source 128-slice CT system, and SF levels were measured at same time. DECT images were acquired using a tube voltage pair of 140 kV and 80 kV or 140 kV and 100 kV, and the three-material decomposition of fat, soft tissue, and iron.[Results]The median age of patients was 52 years (range, 25 to 66), and 8 patients were male. Eight patients with AML, 3 with MDS, and 1 each with ALL, lymphoma, aplastic anemia, Evans syndrome, congenital dyserythropoietic anemia, and chronic renal failure underwent DECT. Nine patients had undergone stem cell transplantation before DECT, and 3 were receiving iron chelate therapy. The total number of units of blood transfused was available in 11 out of 17 patients. The median number of units given was 66 (range, 36 to 150). The median SF level was 2346 ng/ml (range, 569 to 7875). We divided patients into three groups based on SF levels: high >3000 ng/ml, intermediate 1000~3000 ng/ml, low <1000 ng/ml. Five patients were classified into the high SF group (range, 3765-7875 ng/ml), 8 into the intermediate SF group (1645-2916), and 4 into the low SF group (569-1240). Four patients in high, 4 in the intermediate, and 2 in the low SF groups showed diffuse iron deposition in the liver on DECT images. One patient in the high, 2 in the intermediate, and 2 patients in the low SF groups showed focal iron deposition in the left lobe of the liver. On the other hand, two patients in the intermediate SF group did not show iron deposition on DECT images. We then divided patients into two groups based on the number of units of blood transfused: 6 patients were classified into the high unit group (range, 66-150 units), and 5 into the low unit group (36-60). Four patients in the high unit and 3 in the low unit groups showed diffuse iron deposition in the liver on DECT images. Two patients in the high unit and 1 in the low unit groups showed focal iron deposition in the left lobe of the liver. One patient in the low unit group did not show iron deposition.[Conclusion]Discrepancies between SF levels and DECT images indicate that DECT is a useful technique for the accurate evaluation of LIC, and the detection of focal iron deposition in the liver may be useful for optimizing iron chelation therapy. DisclosuresNo relevant conflicts of interest to declare.

Effects of Iron Chelating Therapy and / or HBV - Vaccination on β-Thalassemia Major Patients

Introduction: Thalassemia is a genetic inherited blood disorder in which the body makes abnormal hemoglobin with excessive destruction of red blood cells, which leads to anemia. For many years, hepatitis B virus was a major problem for patients with thalassemia substantially contracted from blood transfusions. The development of effective vaccine has further reduced the magnitude of the problem of hepatitis B. Iron chelators are used to remove excess iron that accumulates due to repeated blood transfusion. Aim of the work: To compare thalassemic patients either have or haven’t HCV with healthy persons as regards biochemical indices taking in consideration effect of vaccination against HBV or not and using iron chelating therapy or not by the studied persons. Methodology: A case control study in which 40 thalassemic, blood transfusion dependent patients were chosen randomly to act as a case group from thalassemic patients attending the VACSERA Company. The cases (40 patients) they were classified to patients having HCV, patients were HBV vaccinated ,other non vaccinated, using iron chelating therapyor don’t use it. Another 10 healthy and non thalassemic persons were chosen randomly among persons attending the same company as a control group to be matched with the case group. Results: 50% of studied thalassemic patients had HCV seropositivity. Biochemical blood indices which were found to be significantly elevated among thalassemic patients than controls were ALT, AST, ALP, GGT and LDH enzymes in addition to serum iron, ferritin and globulin mostly in HBV non-vaccinated and iron chelating therapy non dependent patients while, other biochemical indices which were significantly decreased among thalassemic patients compared to controls included: total cholesterol, total protein, albumin and albumin/globulin ratio mostly in HBV vaccinated and iron chelating non dependent patients. Conclusion: this study illustrated the effectiveness of iron chelators agentsand the importance of vaccination for reduction of morbidity and mortality.

Modification of CYP2E1 and CYP3A4 activities in haemoglobin E-beta thalassemia patients

Thalassemia disease is a genetic haemoglobinopathy usually associated with an iron overload and some degree of organ impairment. The impact of the disease on the drug metabolising enzyme cytochrome P450 (CYP) is not known. CYP2E1 and CYP3A4 are responsible for the metabolism of a large number of drugs and changes in their activities may have clinical consequences. Haemoglobin E-beta thalassemia paediatric, blood transfusion-dependent patients apparently without complications (n = 35) and healthy controls (n = 42) were recruited in this study. The ratios of plasma 6-hydroxychlorzoxazone to chlorzoxazone, and urinary 6-beta-hydroxycortisol (6beta-OHF) to cortisol were used as indices for CYP2E1 and CYP3A4 activities, respectively. Blood and plasma samples were assayed for parameters of clinical biochemistry, oxidants and antioxidants. There were significant increases in serum iron, protein carbonyl and lipid peroxidation in thalassemia patients, whereas there was a decrease in blood glutathione, but unchanged plasma nitric oxide metabolites. CYP2E1 activity in the patients was unchanged; however, when the patients were stratified by splenectomy status, CYP2E1 activity was increased in non-splenectomised patients in comparison with the controls and splenectomised subjects. On the other hand, 6beta-OHF/cortisol ratios increased markedly in patients associated with depressed growth hormone levels. There were no correlations between CYP2E1 activity and oxidant stress or antioxidant parameters. This report is the first demonstration that thalassemia major is associated with an alteration of CYP2E1 and CYP3A4 activities; this could modify the sensitivity of thalassemia patients to the toxic or therapeutic effects of drugs.

Efficacy of Erythropoietin on Dialysis in Patients with Beta Thalassemia Minor

Background: It is unknown whether chronic erythropoietin (EPO) treatment is able to normalize hemoglobin (Hb) levels and ameliorate cardiac remodeling avoiding blood transfusions in uremic blood transfusion-dependent patients with β-thalassemia minor (β-thal). Methods: In 12 hemodialysis (HD) patients with β-thal, requiring blood transfusions despite EPO therapy, we planned to increase Hb levels up to the target levels (11–12 g/dl) within a one-year period by administering progressively higher doses of EPO (correction phase). We also planned to maintain the Hb target for an additional year (maintenance phase). Results: In the year before the study, patients required 3.3 ± 0.9 units of packed red blood cells. At baseline, the Hb level obtained with an EPO dose of 212 ± 73 U/kg/week i.v. was 8.2 ± 0.8 g/dl. The EPO dose was gradually increased within the first year up to 458 ± 78 U/kg/week at month 12 (correction phase) and then significantly tapered down during the maintenance phase (390 ± 54 U/kg/week at month 24). During the correction phase, the Hb levels markedly increased (11.1 ± 0.3 g/dl at month 12) and did not change in the maintenance phase. No blood transfusion was required throughout the 2 years of follow-up. Left ventricular (LV) mass index progressively decreased from the basal value of 144 ± 12 to 124 ± 11 g/m² in the first year and normalized in all patients at month 24 (109 ± 12 g/m², p < 0.001); this occurred in the absence of any change of LV cavity volume index (<90 ml/m²). Conclusions: In HD transfusion-dependent patients with β-thal, the administration of high EPO dose for 2 years permits the attainment and the maintenance of Hb targets without blood transfusions. This therapeutic approach permits a complete remission of concentric LV hypertrophy without any adverse effects on the vascular system.

Molecular Study and Prenatal Diagnosis of Thalassemia in Taiwan

The carrier rates of alpha-thal 1, alpha-thal 2, and beta-thal are 4%, 1% and 1% respectively. If a couple are both the carriers of alpha-thal 1, 25% of the offspring will be affected with homozygous alpha-thal 1, which will result in Hb Bart’s hydrops fetalis and cause various maternal complications. If a couple are both the carriers of beta-thal, 25% of the offspring will be affected with beta-thal major and the patients need life-ling blood transfusion, iron chelation therapy and bone marrow transplantation. This report summarizes our results on thal study in Taiwan. With southern hybridization and various polymerase chain reaction (PCR) techniques, the molecular basis of 832 Chinese Hans with alpha-thal 1 has been characterized. 95.4% had Southeast Asian (SEA) deletion, 2.2% had Thailand (Thai) deletion, and 2.4% had Constant Spring, Quong Sze, or other mutations. 1342 blood samples from five major aboriginal groups, comprising 522 of the Ami, 246 of the Bunun, 227 of the Atayal, 214 of the Paiwan, and 133 of the Yami, were collected for thal study. In the Ami, 43 (8.2%) were alpha-thal 1 carriers (33 with Thai, 9 with SEA, and one with Phil deletion), 43 (8.2%) were alpha-thal 2 carries, one had Hb H disease, and 4 (0.8%) were beta-thal carriers (one each with nonsense 17 and -2 A-G mutation). In the Bunun, one (0.2%) had Thai deletion and 2 (0.4%) were alpha-thal 2 carriers. In the Atayal, one (0.2%) had Phil deletion. In the Paiwan, 7 (3.3%) had alpha-thal 1 (4 with SEA and 3 with Thai deletion) and one (0.5%) had alpha-thal 2. In the Yami, none had either alpha- or beta-thal. In prenatal diagnosis, 624 fetuses at risk for homozygous alpha-thal 1 or severe Hb H disease were studied. Chorionic villus sampling was mainly used to obtain fetal tissue. 170 fetuses were diagnosed as affected. For molecular characterization of beta-thal, 186 unrelated blood-transfusion dependent patients were studied. Southern hybridization, allele-specific oligonucleotide (ASO) hybridization, heteroduplex formation, amplification refractory mutation system (ARMS), artificially created restriction site (ACRS) analysis and direct DNA sequencing have been used. The four most common mutations were IVS-II 654 (C-T)(38.7%), codon 41/42 (deletion of TCTT) (36.6%), -28 (A-G) (6.9%), and nonsense 17 (A-T) (2.8%) mutations. The other mutations detected were A-gamma beta deletion, codon 1 (T-G), codon 27/28 (insertioin of C), Hb E, and codon 71/72 (insertion of A) mutations. The mutations are still unknown in 19 genes. In prenatal diagnosis, 170 fetuses at risk of beta-thal major were studied and 39 fetuses were diagnosed as affected. Three years ago, the Department of Health (DOH) of Taiwan has set up a nation-wide thal screening program. All the pregnant women are encouraged to participate. Couples both with microcytosis are referred to several centers for genotype confirmation and prenatal diagnosis if needed. DOH subsidizes NT 3000 for genotype confirmation of the couple and NT 2000 for prenatal diagnosis. The screening program seems to be running smoothly.