Razi Journal of Medical Sciences

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Background: Identification of the factors influencing short term and long term survival for patients is one of the important concerns in medical society. With determining the risk factors and also health factors of a disease, valuable information can be obtained. In some cases, such as high rate of censoring, the use of standard survival models for analysis may cause to loss in some of information. The purpose of this study was to apply the cure models in identifying the risk and health factors of patients’ thalassemia major.

 

 Methods: The data of this historical cohort study with 296 patients with thalassemia major, were gathered during 1994- 2013 in Zafar Clinic in Tehran. Cure models were used for survival analysis of these patients and determination of its risk factors. Data were analyzed using Stata v.12 and R3.0.2 software.

 

Results: Mean (±SD) of patients’ age was 29.11 ± 0.47year. Of the patients, 51.7% were women. . The minimum value of Akaike information criterion was for cure non mixture log – normal model with logit link function.

 

Conclusion: According to the results, the cure non mixture log – normal models is chosen as the best model for identification the risk factors of thalassemia major disease.

 

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Background: Beta-thalassemia major is a very severe blood disease, its Clinical signs are premature and appear from 3 to 6 months of age. It is one of the most common monogenic diseases in the world and in Iran, and if it is not diagnosed and treated during the first years of life, it will lead to death. In this study, to check the factors affecting the survival of patients with beta-thalassemia major the semi-parametric Cox and parametric Weibull models were used and their efficiency was compared.
Methods: In this longitudinal retrospective study, using the available sampling method, data from 300 patients with beta-thalassemia major referred to Zafar's thalassemia clinic in Tehran during the years 1994 to 2017 were used. To compare the models and select the best model, we used the Akaike information criterion (AIC). Data analysis was performed using Stata software version 13.
Results: The values of the Akaike information criterion (AIC) for semi-parametric Cox model and the parametric Weibull model were 277.600 and 107.211, respectively. Based on both Cox and Weibull models, the variables of blood transfusion, age of desferal and iron deposition in the liver were significantly associated with the survival of patients. Based on Akaike's criterion, the efficiency of the Weibull model was better than the Cox model.
Conclusion: Considering the obtained values, the parametric Weibull model is more efficient than the semi-parametric Cox model for survival analysis of patients with beta-thalassemia major.

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Background: Beta thalassemia is a disorder that is due to the reduction or lack of production of the beta hemoglobin chain, and its inheritance is in the form of autosomal ressesive. The previous program was the prevention of beta thalassemia from the Eighties in Iran and was accompanied by a significant reduction of birthrate with severe and moderate form of disease, and by improving; the cost-effectiveness of the new program was recently in the executive country. Because the main goal of both programs is reducing birthrate, therefore, it was decided to compare these two programs in terms of affected newborn identification power.
Methods: This cross-sectional study was conducted using simple random sampling. Referring to the archive section of Ali-Asghar Children Hospital, the number 72 of patients with thalassemia (36 thalassemia major and 36 thalassemia intermadia) were extracted. In all cases, there was a complete blood count test and hemoglobin electrophoresis for parents of children with the required data including blood markers (RBC, HB, MCV, MCH, HbA2), and age and gender index of parents in the checklist and analyzed statistically by SPSS 20 and SAS 9.1 softwares.
Results: Based on detection type (Thalassemia major or Intermedia) and using new screening protocol and determining the risk of thalassemia, 7.89% of couples in patients with thalassemia major and 100% of couples were considered in the group of patients with thalassemia. Also, based on the usual protocol of risky group, 6.96% of couples in patients with thalassemia major and 100% of couples were enrolled in the group of patients with the Intermedia. In addition, 7.89% were classified for thalassemia major and 100% in Intermedia, without using hemoglobin. Considering the normal ranges of RBC between 4 and 5.5 for children, the cases of anemia had been reported in 5.67%. In addition, in terms of normal range of RBC between 4.7 to 6.1, the cases of anemia in male and female were estimated to be 5.37% and 20%, respectively. In addition, RBCs higher than normal ones were 5.47% and 5.22% respectively. In other words, in men, only 15% and women 5.57% were exposed to normal.
Conclusion: According to the results that were obtained in this study, the use of primary protocols due to the cheaper and more accurate detection of high risk cases for beta-thalassemia are also preferred. In addition, due to the high prevalence of hematologic disorders, especially anemia in these individuals, the implementation of the Protocol for the elimination of anemia in these individuals is strongly recommended according to the national Protocol.

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Background and Aims: Prenatal diagnose (PND) in carrier β-Thalassemia couples is very important. PND of Thalassemia, is a part of national prevention program. In all over the world, conventional prenatal diagnosis of Thalassemia is CVS or AC. These methods are invasive, we want to experience the new NIPD for β-Thalassemia with this hypothesis that 50% of invasive PND removed.
 Non invasive prenatal diagnose of Thalassemia is possible via detection of paternally mutation in maternal plasma. The history of fetal DNA (fDNA) in maternal circulation dates back to the 1948 by Mandel and Metais. The discovery of fDNA in maternal plasma and serum has opened up new opportunities in non invasive prenatal diagnosis (NIPD). This method has been shown to be potentially useful for prenatal diagnosis of certain neurological disorders, fetal sex determination, sex – linked disease, RhD genotyping, and aneuploidies. Fetal DNA originated from trophoblast cells in the placenta, and it detectable very soon during pregnancy, it only comprised about 3 to 6% (although up to 10% has been reported). Fetal DNA fragments are small in size (< 313 bp). It present during pregnancy until birth. They are rapidly cleared within 2 hours after delivery.
Taq- Man is one of the first methods introduced for Real-time sequence detection. A Taq-Man reaction involves 2 specific primers and probes. Each probe is complimentary to one of the alleles of a mutation and each of them is labeled with a different fluorophore. This method exploits the 5' endonucleolytic activity of Taq DNA polymerase to slice an oligonucleotide probe through PCR amplification thus generating a detectable signal.The aim of this study was non invasive prenatal diagnosis of β-Thalassemia and determination the sensitivity and specificity of this technique for detection paternally inherited IVS II-I (GA) mutation in fetus.
Methods: Subjects were selected following ethical approval of study protocol from the ethical committee of Mazandaran University of Medical Science, Iran. The sample size of this study calculated according the related studies . Fifty Iranian pregnant women enrolled in this study, they were at risk of having fetuses with β-Thalassemia major. They attended in prenatal diagnosis of Thalassemia center (at 11-14 weeks of pregnancy, before performance of CVS), between May 2014 and Feb 2015, in Sari, Iran. All women assigned informed consent form. We focused exclusively on couples in which the father was carrier for IVS II-I GàA mutation and the mother had been genotyped to carry another β-globin gene mutation, (Of course, the mutation in each subject had been determined previously by conventional PCR and ARMS method, and followed by agarose gel electrophoresis). Also, 10 milliliters of maternal blood from each pregnant woman were collected into EDTA-containing tube. Maternal plasma was separated from cells by high –speed centrifugation (1900 g or 3000 rpm), and it stored at – 80 0 c until analysis. We were masked as to the identity the samples with numerical coding system, so, these were examined in a blinded manner. We designed two specific primers for IVSII-1(GàA) mutation (Accession No.141900) (www.hgvs.org), on the HBB DNA sequences (NCBI Reference Sequence: NG_000007.3 3 (Homo sapiens beta globin region (HBB)), the oligonucleotid primers of this study were; forward primer: 5'TCACCTGGACAACCTCAAG3', and reverse primer: 5'CCCATTCTAAACTGTACCC3'. The primers pair was optimized to particularly amplify a 200-bp segment of exon 2 of the human β-globin gene. Of course we consider the locus of maternal mutation in β-globin gene that doesn't interfere to our PCR product. Also, for design the Taq-Man probes, we refer in Taq- Man design site (primer digital.com/fast PCR/m15.html FAST PCR manual). The characterics of probes were; GC content 45-65%, no dimer with primer, high Tm 60-65 oc, probe length 18-30 bp, and Tm of primer was 8-10 oc higher than Tm primer. Also, the kind of quencher and reporter were matched with kind of quencher and reporter in the kit. Probe for wild allele was; FAM-AGAACTTCAGGGTGAGTCTATGGG-BHQ1, and the Probe for mutant allele was; VIC-AGAACTTCAGGATGAGTCTATGGG-BHQ1. Circulatory DNA was extracted from 200 µl of peripheral blood plasma with QIAamp DNA mini and blood kit according to the manufacturer's instructions. DNA was eluted in a final volume of 200 µl AE buffer and stored at -20 0 c until used for additional analyzing. Also, the genomic DNA concentration was measured by NanoDrope system and electrophoresis. For detection of paternally inherited mutation Taq-Man Real time PCR was used and the result of Taq- man method compared with result of chorionic vilus sampling method (CVS) as a gold standard.
Results: The clinical performance of Taq-Man test was assayed by comparing with routine test. Included in this study were 50 pregnant mothers. In Taq-Man method 38 samples cited in scatter analysis graph as a negative control, because the concentration of DNA was very low and this method didn’t have enough sensitivity and specificity for detection, so, the final analysis for assessment of this method was done on 12 samples. We reported 5 cases true positive, 4 cases true negative, 2 cases false negative and 1 cases false positive. Also the sensitivity of Taq-Man was 71.43% with CI95% (60%-83.43%), and specificity 66.67% with CI 95% (54.67%-78.67%). LR+ and LR- were 2.15 and 0.5 respectively. Also NPV and PPN of Taq-Man test were 66.67%, 83.4% respectively.
Conclusion: Non invasive prenatal diagnose can be detect paternally inherited mutation of thalassemia.