Antonietta Zappu

Bannayan-Riley-Ruvalcaba syndrome with reactive nodular lymphoid hyperplasia and autism and a PTEN mutation.

Loredana Boccone,* Valentina Dessı̀, Antonietta Zappu, Silvia Piga, Maria Bonaria Piludu, Marco Rais, Carlo Massidda, Stefano De Virgiliis, Antonio Cao, and Georgios Loudianos Ospedale Regionale Microcitemie, Cagliari, Italy Dipartimento di Scienze Biomediche e Biotecnologie, USC, Italy Istituto di Neurogenetica e Neurofarmacologia, CNR-Cagliari, Italy U.O. Anatomia Patologica, Ospedale Businco, Cagliari, Italy Chirurgia Oncologica, Ospedale Businco, Cagliari, Italy

Efficient strategy for molecular diagnosis of wilson disease in the Sardinian population

Wilson disease (WD) is an autosomal recessive disorder of copper transport resulting from the defective function of a copper-transporting ATPase (ATP7B) (1)(2)(3). More than 200 disease-causing mutations have been identified (4). In the Sardinian population, WD has an incidence of ∼1 in 7000 live births (5). Using single-strand conformation polymorphism (SSCP) and sequencing methods for mutation analysis, we have characterized 92% of the chromosomes analyzed and identified 16 different WD-causing mutations, 6 of which (−441/−427del, 213–214delAT, 1512–1513insT, R778W, 2463delC, and V1146M) are relatively common and account for 85% of chromosomes (6). On the basis of these data, we developed a reverse dot-blot (RDB) method as a practical solution to mutation screening in this population. DNA samples from Sardinian WD patients carrying different combinations of the most common mutations (−441/−427del, 2463delC, V1146M, 213–214delAT, 1512–1513insT, and R778W) were used as controls. Our aim was to obtain the same PCR conditions for all six pairs of primers that were used to amplify the regions containing the six most common mutations. We therefore designed primers with an identical melting temperatures and tested their specificity first in single and then in multiplex PCRs. We also wanted to obtain relatively equal yields for all PCR products to obtain comparable signals using the RDB method. We therefore tested different concentrations for each pair of primers and finally established primer concentrations that allowed us to obtain approximately …

The homozygosity index (HI) approach reveals high allele frequency for Wilson disease in the Sardinian population

Wilson disease (WD) is an autosomal recessive disorder resulting in pathological progressive copper accumulation in liver and other tissues. The worldwide prevalence (P) is about 30/million, while in Sardinia it is in the order of 1/10 000. However, all of these estimates are likely to suffer from an underdiagnosis bias. Indeed, a recent molecular neonatal screening in Sardinia reported a WD prevalence of 1:2707. In this study, we used a new approach that makes it possible to estimate the allelic frequency (q) of an autosomal recessive disorder if one knows the proportion between homozygous and compound heterozygous patients (the homozygosity index or HI) and the inbreeding coefficient (F) in a sample of affected individuals. We applied the method to a set of 178 Sardinian individuals (3 of whom born to consanguineous parents), each with a clinical and molecular diagnosis of WD. Taking into account the geographical provenance of the parents of every patient within Sardinia (to make F computation more precise), we obtained a q=0.0191 (F=7.8 × 10−4, HI=0.476) and a corresponding prevalence P=1:2732. This result confirms that the prevalence of WD is largely underestimated in Sardinia. On the other hand, the general reliability and applicability of the HI approach to other autosomal recessive disorders is confirmed, especially if one is interested in the genetic epidemiology of populations with high frequency of consanguineous marriages.

The canine copper toxicosis gene MURR1 is not implicated in the pathogenesis of Wilson disease

BackgroundIt has recently been demonstrated that the Wilson disease (WD) protein directly interacts with the human homolog of the MURR1 protein in vitro and in vivo, and that this interaction is specific for the copper transporter. The aim of the present study was to clarify the role of MURR1 in the pathogenesis of WD as well as in other WD-like disorders of hepatic copper metabolism of unknown origin.MethodsUsing the single-strand conformation polymorphism (SSCP) method followed by sequencing, we analyzed the 5′ untranslated region (UTR) and three exons of the MURR1 gene in three groups of patients: 19 wd patients in whom no mutations were detected in the ATP7B gene, 53 wd patients in whom only one mutation in the ATP7B gene was found, and 34 patients in whom clinical and laboratory data suggested a WD-like disorder of hepatic copper metabolism of unknown origin.ResultsWe  detected in these patients six rare nucleotide substitutions, namely one splice-site consensus sequence and one missense and four silent nucleotide substitutions. All substitutions except one were found in the heterozygous state. No difference in the frequencies of the various substitutions was observed between patients and controls.ConclusionsThese data suggest that the MURR1 gene and its protein product are unlikely to play a primary role in the pathogenesis of Wilson disease. More extensive studies with larger numbers of clinically homogeneous patients should be carried out to establish whether nucleotide alterations in the MURR1 gene may have a role in causing WD or WD-like disorders or act as modifying factors in the phenotype variability in WD.

Mutation analysis of the ATP7B gene in a new group of Wilson's disease patients: Contribution to diagnosis.

Wilsons disease (WD), an autosomal recessive disorder of copper transport with a broad range of genotypic and phenotypic characteristics, results from mutations in the ATP7B gene. Herein we report the results of mutation analysis of the ATP7B gene in a group of 118 Wilson disease families (236 chromosomes) prevalently of Italian origin. Using DNA sequencing we identified 83 disease-causing mutations. Eleven were novel, while twenty one already described mutations were identified in new populations in this study. In particular, mutation analysis of 13 families of Romanian origin showed a high prevalence of the p.H1069Q mutation (50%). Detection of new mutations in the ATP7B gene in new populations increases our capability of molecular analysis that is essential for early diagnosis and treatment of WD.

RNA analysis of consensus sequence splicing mutations: implications for the diagnosis of Wilson disease.

Wilson disease (WD) is an autosomal recessive disorder caused by a defective function of the copper-transporting ATP7B protein. This results in progressive copper overload and consequent liver, brain, and kidney damage. Approximately 300 WD-causing mutations have been described to date. Missense mutations are largely prevalent, while splice-site mutations are rarer. Of these, only a minority are detected in splicing consensus sequences. Further, few splicing mutations have been studied at the RNA level. In this study we report the RNA molecular characterization of three consensus splice-site mutations identified by DNA analysis in WD patients. One of them, c.51 + 4 A --> T, resides in the consensus sequence of the donor splice site of intron 1; the second, c. 2121 + 3 A --> G, occurred in position + 3 of intron 7; and the c.2447 + 5 G --> A is localized in the consensus sequence of the donor splice site of intron 9. Analysis revealed predominantly abnormal splicing in the samples carrying mutations compared to the normal controls. These results strongly suggest that consensus sequence splice-site mutations result in disease by interfering with the production of the normal WD protein. Our data contribute to understanding the mutational spectrum that affect splicing and improve our capability in WD diagnosis.

Development of TaqMan allelic specific discrimination assay for detection of the most common Sardinian Wilson's disease mutations. Implications for genetic screening

Wilsons disease (WD) is an autosomal recessive disorder caused by a defective function of the copper transporting ATP7B protein. Analysis of ATP7B gene in the Sardinian population revealed the presence of six common mutations that together account for 85% of WD chromosomes. We have developed an automated approach for the detection of these 6 common Sardinian mutations based on TaqMan technology. Ten DNA samples of WD patients carrying different combinations of the six most common Sardinian mutations and normal controls previously analysed were used in triplicate to set up the allelic discrimination assays. The system was validated in 96 samples obtained from WD patients carrying different combinations of the most common mutations under investigation. The results showed that allelic discrimination is a valid method that could be used for efficient diagnosis of single cases but also for a mass screening.

Deferasirox for cardiac siderosis in β-thalassaemia major: a multicentre, open label, prospective study

10 per anti-IIa level draw. Assuming an average weight patient, the estimated cost of argatroban in patients in the aPTT cohort was estimated to be

Causes of hospital admission in children and adults with transfusion-dependent thalassemia in Sardinia, 2000–2015

7628 US dollars compared to