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M Tzetis

Publications and source records attributed to M Tzetis.

29 records · Page 2Linked to original sources
PubMed1997

Prenatal diagnosis of the thalassaemia syndromes by rapid DNA analytical methods.

Prenatal diagnostic strategies applied today are based mainly on polymerase chain reaction (PCR) analytical protocols. In Greece a wide range of mutations underlie the thalassaemic haemoglobinopathies, and consequently a variety of PCR-based methods are required to facilitate diagnosis of all potential abnormal genotypes. PCR protocols include those which are relatively simple and others that are technically challenging, but very few have been designed for high through-put clinical diagnostics. Over a period of 18 months we carried out prenatal diagnosis of 147 pregnancies (150 fetal samples) at risk for a wide range of haemoglobinopathies. This involved the precise characterization of parental genotypes and the subsequent analysis of fetal DNA samples. In this series, 18 different mutations in the alpha- or beta-globin clusters were identified. For the characterization of these mutations, five PCR-based protocols were selected: denaturing gradient gel electrophoresis (DGGE), amplification refractory mutation system (ARMS) PCR, restriction endonuclease analysis of PCR fragments, oligonucleotide hybridization and 'gap' PCR for detection of deletions. To avoid spurious diagnosis due to contamination of fetal samples, two additional methods were used to genotype polymorphic variable nucleotide tandem repeat (VNTR) regions of the genome in parental and fetal samples. Through analysis of the results we assess the advantages and drawbacks of the selected PCR-based protocols for providing routine clinical diagnostics.

Alleles↗
PubMed1995

Mutation analysis of ten exons of the CFTR gene in Greek cystic fibrosis patients: characterization of 74.5% of CF alleles including one novel mutation.

To initiate the complete characterization of mutations in the CFTR gene in Greek cystic fibrosis (CF) patients, we screened 184 patients for six relatively common mutations (delta F 508, G542X, G551D, 621 + 1 G-->T, N1303K, W1282X) using allele-specific hybridization and, in addition, analyzed exons 4, 5, 7, 8, 10, 11, 17b, 19, 20 and 21 using the method of denaturing gradient gel electrophoresis (DGGE). Six mutations accounted for 65.9% of the CF alleles in Greek patients, of which the delta F 508 mutation had a frequency of 52.7%. A further 15 previously described mutations accounted for another 8.3% CF alleles and one previously undescribed mutation (3272-4A-->G) was found in one chromosome. The W1282X mutation was not detected at all. Thus, so far, we have identified 21 mutations in the CFTR gene in Greek CF patients, accounting for 74.5% of the CF alleles.

Alleles↗
PubMed1994

The molecular basis of normal HbA2 (type 2) beta-thalassemia in Greece.

Heterozygotes for beta-thalassemia usually have raised levels of HbA2, but in Greece about 5% of beta-thalassemia carriers are observed to have normal or borderline levels. It is postulated that such cases have mild beta+ thalassemia mutations or coinheritance of delta-thalassemia. We selected 18 heterozygotes with the hematological phenotype of normal HbA2 (type 2) beta thalassemia who were negative for the delta beta Corfu mutation, and screened them for previously defined Mediterranean beta-thalassemia and delta-thalassemia mutations. The coinheritance of beta and delta-thalassemia was demonstrated in four cases with the following genotypes: in cis beta+ IVSII -n745/delta+ 27, beta 0NS39/delta 059(-A), beta+ IVSI-n110/delta 059(-A) and in trans beta+ IVSI-n6 and delta+ 27. A further nine heterozygotes had mild beta(+)-thalassemia mutations (eight with the beta+ IVSI-n6 mutation, one with the beta+ polyA (A-->G) mutation). In four heterozygotes with severe beta-thalassemia chromosomes (2 beta+ IVSI-n110, 1 beta 0 FSC-6, 1 beta 0 IVSI-n1) no known delta-thalassemia mutations were observed. One case had a delta beta deletion chromosome. These results indicate that the hematological phenotype of normal HbA2 (type 2) beta-thalassemia in Greece is genetically heterogeneous; it is mainly associated with the delta beta Corfu mutation or coinheritance of beta and delta thalassemia mutations or with very mild beta(+)-thalassemia mutations, mainly beta+ IVSI-n6. In the rare cases with severe beta-thalassemia mutations, the normal levels of HbA2 may be due to coinheritance of as yet undefined delta thalassemia mutations.

Base Sequence↗
PubMed1993

Characterization of nondeletion alpha-thalassemia mutations in the Greek population.

alpha-Thalassemia is usually due to deletions within the alpha-globin gene cluster, leading to loss of function of one (-alpha) or both [-(alpha) or --] alpha-globin genes. Nondeletion mutations (denoted alpha alpha T or alpha T alpha) are less frequent and in Greece are not well defined. We report the analysis of 16 nondeletion alpha-thalassemia chromosomes using a polymerase chain reaction method to amplify specifically the alpha 2-globin gene, which was subsequently screened using ASO hybridization or restriction enzyme analysis for four mutations already characterized in other Mediterranean and Middle Eastern populations. Of the 16 nondeletion chromosomes, nine had the polyadenylation signal mutation (alpha PolyA alpha), two the IVSI 5' pentanucleotide deletion (alpha Hph alpha), two the Hb Icaria mutation (alpha Ic alpha), and one the initiation codon mutation (alpha Nco alpha). In two, the defects are still undefined. These findings show that nondeletion alpha-thalassemia in Greece is heterogeneous and that the most frequent mutation (accounting for > 50%) is the polyadenylation signal mutation, which to date was most commonly found in the Saudi Arabian population.

Alleles↗
PubMed1991

The Corfu delta beta thalassaemia mutation in Greece: haematological phenotype and prevalence.

The Corfu delta beta thalassaemia mutation, a 7.2 kb deletion partially removing the delta-globin gene and a single nucleotide mutation (G----A) at intervening sequence I (IVSI-n5) in the beta-globin gene in cis, was first described in a family from Corfu; the carriers for this mutation had the unusual haematological phenotype of heterozygous beta-thalassaemia with normal levels of HbA2. To investigate the frequency and haematological characteristics of Corfu delta beta thalassaemia in Greece we analysed 25 unrelated normal HbA2 type 2 beta-thalassaemia heterozygotes and their 23 clinically affected offspring. Gene mapping demonstrated that nine (36%) of the 25 normal HbA2 beta-thalassaemia heterozygotes were in fact Corfu delta beta thalassaemia heterozygotes and of the 23 patients, two were Corfu delta beta thalassaemia homozygotes and five compound heterozygotes for Corfu delta beta thalassaemia and another beta-thalassaemia defect. Detailed haematological analysis demonstrated that: the Corfu delta beta thalassaemia mutation does not completely abolish the expression of the beta-globin gene; the HbA2 levels are slightly lower (P less than 0.01) and the HbF levels slightly higher (P less than 0.01) in Corfu delta beta thalassaemia heterozygotes compared to beta-thalassaemia heterozygotes with the normal HbA2-type 2 phenotype who do not have the Corfu delta beta chromosome.

Chromosome Deletion↗
PubMed1988

Clinical, haematological, and genetic studies of type 2 normal Hb A2 beta thalassaemia.

The clinical and haematological phenotype as well as chain synthesis data were studied in 35 doubly heterozygous patients with either normal Hb A2 and Hb F, type 2 beta thalassaemia and beta (high A2) thalassaemia (26 patients), or type 2 and other rare beta or delta beta variants (nine patients). Patients doubly heterozygous for type 2 and beta zero or delta beta zero thalassaemia variants had no detectable Hb A, indicating that the type 2 normal A2 beta thalassaemia is primarily the result of a beta zero gene. The clinical phenotype varied from severe thalassaemia major to mild thalassaemia intermedia, and was mainly related to the thalassaemia variant with which the type 2 normal A2 beta thalassaemia was combined, and the proportion of Hb A produced in beta + thalassaemia patients. Haematological and chain synthesis data were similar in heterozygotes with type 2 and beta zero or beta + (high A2) thalassaemia. Hb A2 levels were within the normal range (2.3 to 3.6%) though absolute values (Hb A2 per RBC) ranged from low normal (0.5 pg/RBC) to increased levels (1.0 pg/RBC.) The variation of Hb A2, as well as the presence of Hb A2 in a type 2/delta beta high F patient and the complete absence of HbA2 in a homozygous type 2 patient, indicate that there are at least two genotypes of type 2, one beta zero and the other delta beta zero. This has been recently proven by gene mapping studies. For clinicians, routine haematological and family studies are sufficient for the proper treatment and prevention of doubly heterozygous type 2 patients.

Blood Protein Electrophoresis↗
PubMedDate not supplied

Haplotype and mutation analysis in Greek patients with Wilson disease.

In this study, we report the results of haplotype and mutation analysis of the ATP7B gene in Wilson disease (WD) patients of Greek origin. We have analysed 25 WD families and two single patients and characterised 94% of the WD chromosomes investigated. We have found 12 different molecular defects (three frameshifts, two splice site, two nonsense, five missense mutations), four of which are novel. Five of the mutations are widely prevalent accounting for 74% of the WD chromosomes analysed. These results may enable preclinical diagnosis in the large majority of WD patients of Greek descent, thereby improving genetic counselling and disease management.

Adolescent↗
PubMedDate not supplied

Hematologic phenotype of the mutations IVS1-n6 (T-->C), IVS1-n110 (G-->A), and CD39 (C-->T) in carriers of beta-thalassemia in Greece.

The hematologic phenotype was characterized in heterozygotes for three of the most common beta-thalassemia mutations in the Greek population. The study included 17 carriers of beta++ IVS1-n6 (T-->C), 21 carriers of beta+ IVS1-n110 (G-->A), and 17 carriers of beta 0 CD39 (C-->T). The 55 beta-thalassemia heterozygotes were selected from among parents of patients on regular transfusion regimens, and the beta-thalassemia mutation was identified by means of the polymerase chain reaction to amplify the appropriate region of the beta-globin gene and then by allele-specific oligonucleotide hybridization. The assessment of hematologic phenotype included complete blood count and quantitation of hemoglobin HbA2 and HbF and of the globin chain biosynthesis ratio. Comparison and statistical analysis of the hematologic parameters for the three mutations demonstrated no consistent correlation among the three mutations relative to Hb levels, hematocrit, and red cell indices, although heterozygotes for the IVS1-n6 mutation produce red blood cells with slightly higher mean corpuscular volume; significantly lower values of HbA2 (mean, 3.81% +/- 0.62% with four values less than 3.60%) in IVS1-n6 heterozygotes compared with IVS1-n110 heterozygotes (mean, 4.69% +/- 0.48%) and CD39 heterozygotes (mean, 4.75% +/- 0.50%, P < 0.001); and significantly higher HbF levels in CD39 heterozygotes (mean, 2.31% +/- 1.52%) compared with IVS1-n6 heterozygotes (mean, 0.79% +/- 0.45%, P < 0.01) and IVS1-n110 heterozygotes (mean, 1.17% +/- 0.75%, P < 0.01). With respect to the HbA2 levels, the findings are in agreement with previous studies in Mediterranean populations; the slightly higher levels of HbF in CD39 heterozygotes appear to be reported for the first time.

Alleles↗
PubMedDate not supplied

Molecular characterization of homozygous (high HbA2) beta-thalassemia intermedia in Greece.

Homozygous beta-thalassemia is usually characterized by severe anemia requiring regular blood transfusion for survival. For homozygous patients with milder clinical manifestations and no dependence on transfusion therapy, the term thalassemia intermedia is usually applied. Genetic mechanisms that may ameliorate the clinical expression of homozygous beta-thalassemia include coinheritance of alpha-thalassemia, inheritance of mild beta-globin gene mutations, and increased gamma-globin chain production, which may partially compensate for the lack of beta-globin chain synthesis. To identify which of these factors may contribute to the modification of childhood homozygous, high-hemoglobin A2 (HbA2) beta-thalassemia in Greece, the interaction of alpha-thalassemia, types of beta-thalassemia mutations, and the presence of a polymorphic site 5' to the G gamma-globin gene, which has been described as associated with increased gamma-globin chain production in some cases, was assessed. The results were analyzed in light of similar studies in 150 randomly selected, homozygous, high-HbA2 beta-thalassemia patients with the aim of assessing whether thalassemia genotypes can provide information useful for prognosis and/or more appropriate management of homozygous beta-thalassemia patients. The results indicate that, in general, the major factor modifying the clinical expression of homozygous, high-HbA2 beta-thalassemia in Greece is the inheritance of mild beta-thalassemia mutations. Although there is not always a complete correlation of genotype with clinical phenotype, the inheritance of two mild beta-thalassemia alleles results in almost all cases (11 of 12 cases in this study) in thalassemia intermedia phenotype.(ABSTRACT TRUNCATED AT 250 WORDS)

Child↗