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Abstract
All patients were of Egyptian origin and most of them were immigrant from 14 different Egyptian cities situated on the Nile Delta and Upper Egypt. In their results, we revealed that there is no relation between genotyping and phenotyping in the majority of the patients so mutations in both alpha and gamma genes were possibilities to affect the phenotypic character of the disease.
Therefore, this work aimed to study the most common mutation in γ- globin gene in β- thalassemic patients. Many studies tried to link the Xmn1 polymorphism (C-T) at position -158 of γ- globin gene with β- thalassemic patients and its relation with their phenotype.
Up to our knowledge, this study is the first investigation on molecular basis of the γ- globin genes in β- thalassemic patients in Suez Canal region and the second in the Egyptian population (that was done outside Egypt).
Samples were collected from March through August 2006, this study included 32 beta thalassemia patients (21 males and 11 females) belonging to 24 families living in Ismailia and Port- Said, 17 of them (53.1%) were thalassemia major while the rest of them (15/32, 46.9%) were thalassemia intermedia.
Detailed history was recorded for all patients. This included personal history (age, sex and consanguinity), detailed history of the disease (age of diagnosis, 1st transfusion, frequency of transfusion, the presence of any complications and the history of treatment) and family history.
Patients were examined by the members of Hematology Pediatric Clinic for general examination to detect the signs of anemia, presence of thalassemic facies, growth and developmental retardation and abdominal examination to detect liver and splenic enlargement.
The beta thalassemic patients were 21 males (65.6%) and 11 females (34.4%), the ration between males and females 1.9:1. Their age ranged from 5 to 33 years old (mean 14.1 + 5.6 years), most of them were children (10 patients) and adolescent (18 patients) while only four patients reached the adult life.
All the patients were diagnosed during infancy and childhood. The age of diagnosis ranged from 2 months to 4 years old. Most of the patients were diagnosed in the first year of life (18/32, 56.3%) while the rest were in next three years old (14/32, 43.7%). On the other hand, a positive consanguinity was found in 14/32 patients (43.8%) while the rest 18/32 patients (56.2%) were negative consanguinity.
Hb level was measured in all our patients, it ranged from 6 gm% up to 11 gm% (mean 7.8 + 1.5 gm%), in thal major patients the mean was 7.2 + 1.2 gm% while in thal intermedia patients it was 8.4 + 1.5 gm% (P= 0.017). There was no statistically significant difference between male (8.4 + 2.4 gm %) and female (8.5 + 2.5 gm %) β- thalassemia patients. Hb F ranged from 0.8- 45.4% of total Hemoglobin (mean 9.4 + 9.6%); in thal major the mean was 6.8 + 3.7% while in thal intermedia it was 12.4 + 12.9% (Hussein et al., 2005).
Six patients (18.8%) were not receiving any blood transfusion during their whole lives. The majority of patients (17/32, 53.1%) were transfused once every 3-5 weeks, while six patients (18.8%) were transfused every 2-4 months and three patients (9.3%) were transfused more frequently every 2 weeks.
The spleen was normal in eight patients (25%). It was enlarged in eleven patients (34.4%). Thirteen patients were splenectomized (40.6%); the age of splenectomy ranged from 1.5 to 15 years old.
Serum ferritin was normal only in one female patient (3.1%), while it was elevated in the rest 31 patients. In females, the mean was 4123.6 + 2866.8 ng/ ml; while it was lower in males (because of their higher number in the study) it was 3814.3 + 2984.6 ng/ ml.
According to the type of thalassemia, serum ferritin was higher in thalassemia major patients than thalassemia intermedia patients.
Liver status (by clinical examination) was normal in 56.3% of the patients; thalassemic face was noticed in 19 cases (59.4%). Most of the patients (18/32, 56.2%) showed retarded linear growth (> 2 SD for age and sex).
C\T polymorphism at position -158 of γ- globin gene was studied by PCR amplification of the gamma globin promoter fragment (-1227 to +30) followed by Xmn1 restriction enzyme digestion and detection by agarose gel electrophoresis. Lambda plasmid λ was used as a positive control.
Xmn1 polymorphism in Gamma globin gene was found in 2 patients (6.25%); the 1st patient was 21 years old male with thalassemia major, his type of mutation was heterozygous FS5 (-CT)/IVSI:6 and Hb F was 3.5%. The other patient was 17 years old female with thalassemia intermedia, her type of mutation was homozygous IVSII: 1 and Hb F was 10.2%.
from these results we could found that Xmn1 polymorphism in Gamma globin gene was 5.88% of all thalassemia major patients and 6.67% of all thalassemia intermedia, but it was observed that no relationship was found between the presence of this polymorphism and any other criteria like sex, type of mutation in β- globin gene, Hb F percentage, splenic & liver status or consanguinity.
In conclusions: It was observed that C\T polymorphism at position -158 of γ- globin gene is present in some patients with beta-thalassemia major and intermedia and it does not seem, in this study, to influence the phenotyping or Hb F content.