Prenatal diagnosis of haemoglobinopathies: our experience of 523 cases

Abstract Background: We performed counselling for prenatal diagnosis (PD) of haemoglobinopathies in 372 couples. Thirty-four out of 372 (9.1%) did not undergo PD: six due to spontaneous abortion; nine because it was too difficult to make a decision if PD was positive; 18 because counselling excluded the carrier status of one or both parents; and one because parental mutations were mild. Methods: Eleven out of 338 (3.3%) couples underwent PD because they had a thalassaemic child; 106 (31.4%) were found to be at high risk during pre-conceptional screening; 221 (65.4%) because of familiarity. Of 523 PDs in 486 (92.9%), including six dichorionic twin pregnancies, PD was performed on DNA from chorionic villi (CV), and in 37 from amniocytes (7.1%). In 1/523 cases, PD was not completed because DNA from CV was not sufficient; in two cases single tandem repeat analysis revealed maternal contamination of foetal DNA; in 7/522 (1.3%) cases PD revealed non-paternity. In 435/522 (83.3%) cases, PD was performed using reverse dot-blot and ARMS; 34/522 (6.5%) required sequencing. In 53/522 (10.2%) cases it was necessary to test globin loci for large rearrangements. Results: One hundred and twenty out of 522 (23.0%) PDs revealed an affected foetus. In all but two cases the couple interrupted pregnancy. In the six twin pregnancies PD revealed a normal and a carrier foetus (two cases), carrier status in both foetuses (two cases) and a carrier and an affected foetus (two cases). In these latter cases the couple planned selective interruption. Conclusions: Our PD procedure is successful and reliable, and is useful in high-risk areas characterised by molecular heterogeneity

Defective mRNA levels are responsible for a beta-thalassemia phenotype associated with Hb Federico II, a novel haemoglobin variant (beta 106 (G8) Leu->Val).

This study provides the first experimental evidence that a single nucleotide mutation within the coding region of the β-globin gene affects mRNA expression levels and causes a β-thalassemic defect. Furthermore, our data suggest that other regions besides the 3′UTR, whose role in constitutively regulation of this mechanism has been recently identified, may contribute to the stabilization of β-globin mRNA and could, therefore, help to characterize the molecular basis of thalassemic hemoglobinopathies

Role of the Cold Shock Domain Protein A in the transcriptional regulation of gamma-globin gene expression.

International audienceImpaired switching from fetal haemoglobin (HbF) to adult globin gene expression leads to hereditary persistence of fetal haemoglobin (HPFH) in adult life. This is of prime interest because elevated HbF levels ameliorate beta-thalassaemia and sickle cell anaemia. Fetal haemoglobin levels are regulated by complex mechanisms involving factors linked or not to the beta-globin gene locus. To search for factors putatively involved in gamma-globin gene expression, we examined the reticulocyte transcriptome of three siblings who had different HbF levels and different degrees of beta-thalassaemia severity although they had the same alpha- and beta-globin gene cluster genotypes. By mRNA differential display we isolated the cDNA coding for the cold shock domain protein A (CSDA), also known as dbpA, previously reported to interact in vitro with the gamma-globin gene promoter. Expression studies performed in K562 and in primary erythroid cells showed an inverse relationship between gamma-globin and CSDA expression levels. Functional studies performed by Chromatin Immunoprecipitation and reporter gene assays in K562 cells demonstrated that CSDA is able to bind the promoter of the gamma-globin gene and to suppress its expression. Therefore, our study demonstrates that CSDA is a trans-acting repressor factor of gamma-globin gene expression and contributes to modulate the HPFH phenotype