P Sammarco

Epidemiological study of nonsyndromic hearing loss in Sicilian newborns

Deafness is caused by a variety of facts, genetic and environmental. Regarding the acquired causes, deafness can be the consequence of prenatal infections, acoustic or cerebral trauma, and the use of ototoxic drugs. Deafness can be the only manifestation (nonsyndromic forms) or it may occur together with other phenotypic findings (syndromic forms). The majority of nonsyndromicdeafness has a genetic basis [Van Camp et al., 1997]. In recent years, deafness and hearing loss have assumed a clinical importance in the study of congenital disorders [Morton et al., 1991]. The clinical interest for hearing loss is supported by the social impact that this disorder has; if not treated, delays in the development of language and learning skills will occur [Yoshinaga-Itano et al., 1998]. In the absence of newborn screening, hearing loss might not be noticed by parents, teachers or paediatricians until the child begins to have difficulties at speaking and learning, sometimes as late as age 2 or 3 years. In genetic nonsyndromic hearing loss (NSHL) the inheritance pattern is autosomal recessive (80%), autosomal dominant (17%), X-linked (2–3%), and mitochondrial (<1%) [Snoeckx et al., 2005]. Overall NSHL is very heterogeneous; about 100 loci have been related to NSHL and 37 genes encode for proteins which have a role in inner ear physiology [Snoeckx et al., 2005].However, variants of one gene, GJB2 (gap junction protein, beta 2, OMIM#220290) account for up of 50% of NSHL in many populations [Kenneson et al., 2002]. TheGJB2 gene is expressed in a variety of cells and tissues. It encodes connexin 26 protein (CX26), one member of a great number proteins family which are involved in the formation of the splice gap (gap-junctions) that allow the direct transfer of small ionic molecules between adjacent cells. In the cochleaCX26-containing gap junctions areproposed to maintain Kþ homeostasis between outer hair cells and endolymphatic space during the auditory transduction [Wangemann, 2002]. Recently, it has been know that the intracellular transduction of second messenger inositol triphosphate (IP3) is also essential for the perception of sound [Beltramello et al., 2005]. The GJB2 gene is localized on chromosome 13 (13q11-q12) [Guilford et al., 1994], and it contains two exons with only the second is translated [Griffith et al., 2000]. Many NSHL-causing mutations of GJB2 have been reported [Murgia et al., 1999] and they have been stored in world wide gene mutations database http:// www.crs.es/deafness. Based on published data, these GJB2 variant have been classified as truncating and nontruncating mutations [Snoeckx et al., 2005]. The group of truncating contains nonsense mutations, deletions, insertions, and duplications that create an anticipated stop codon; the splice site mutation (IVS1þ 1 G!A) is classified as truncating [Snoeckx et al., 2005]. The group of nontruncating contains amino acid substitutions and one in frame deletion [Snoeckx et al., 2005] which could have severe phenotypic consequences because for some amino acid substitutions, CX26 function could be lost. The most common truncating mutations are the small deletions 35delG, 167delT, 235delC [Snoeckx et al., 2005], the splice site mutation IVS1þ 1 G!A [Denoyelle et al., 1999] however, the prevalence

Distribution and phenotype of GJB2 mutations in 102 Sicilian patients with congenital non syndromic sensorineural hearing loss

Abstract Objective: To evaluate the frequency of GJB2 mutations and their correlation with phenotype in Sicilian non-syndromic sensorineural hearing loss (NSHL) patients. Design: Sequencing of the coding region, basal promoter, exon 1, and donor splice site of the GJB2 gene; screening for the presence of the two common GJB6 deletions. Study sample: A cohort of 102 Sicilian NSHL patients. Results: Fifteen different mutations in GJB2 and seventeen different genotypes were detected. No GJB6 mutations were found. The hearing impairment was profound in the 64.72% of probands (mean PTA0.25–4 kHz of 88.82 ± 26.52 dB HL). A total of 81.37% of patients harboured at least one c.35delG allele; c.167delT and c.-23 + 1G> A were identified in 10.78% and the 9.8% of patients respectively; c.35delG homozygotes presented more severe hearing impairment (75.59% of profound hearing loss) and a higher mean PTA0.25–4 kHz (96.79 ± 21.11 dB HL) with respect to c.35delG/non-c.35delG and c.35delG/Wt patients (P < 0.05). Conclusions: This work underlines the role of c.35delG, c.167delT and c.-23 + 1G> A as the most frequent causes of NSHL in Sicily. The c.35delG frequency found is similar to those reported in other populations of the Mediterranean area. The analysis of genetic and audiologic data confirmed a variability in the phenotype associated to a single genotype.

Persistent jaundice in an infant with homozygous beta thalassemia due to co-inherited Crigler-Najjar syndrome.

Clinically apparent jaundice is unusual in patients with β‐thalassemia major. Co‐inheritance of Gilbert syndrome has been reported to cause hyperbilirubinemia in these subjects. Crigler–Najjar syndrome is another rare disorder of bilirubin metabolism caused by mutation in the gene coding the enzyme UGT1A1. We report a patient of β‐thalassemia major who presented with persistent jaundice due to co‐inherited Crigler–Najjar syndrome type 2 secondary to a novel mutation in UGT1A1 gene [homozygous base substitution at position 362 (GGT>AGT) in exon 3]. Pediatr Blood Cancer 2010;54:627–628.

Glucose 6-phosphate dehydrogenase Palermo R257M: a novel variant associated with chronic non-spherocytic haemolytic anaemia.

Glucose 6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzyme defect, present in over 400 million people worldwide (Frank, 2005; Cappellini & Fiorelli, 2008). It is an X-linked hereditary genetic defect caused by mutations in the G6PD gene. About 140 variants have been described worldwide (Cappellini & Fiorelli, 2008). These variants can be grouped into five classes according to their level of residual enzyme activity and of phenotypes (Betke et al, 1967; Fiorelli et al, 2000; Cappellini & Fiorelli, 2008). G6PD mutations, described as Class I variants, include the most severe form of G6PD deficiency and lead to chronic nonspherocytic haemolytic anaemia (CNSHA) (Fiorelli et al, 2000). The best known mutations leading to CNSHA are clustered in exon 10, which encodes the presumptive dimerization site, although other mutations have been described in exons 7, 8 and other regions of the gene (Vulliamy et al, 1993; Costa et al, 2000). CNSHA is characterized by a chronic haemolytic anaemia with intermittent exacerbations of acute intravascular haemolysis after exposure to oxidant stress, often requiring blood transfusion and leading to jaundice, hyperbilirubinaemia, moderate splenomegaly and reticulocytosis. Gallstones are also frequently detected. However, some variants associated with CNSHA show a haemoglobin concentration within the normal range, with haemolysis well compensated (Luzzatto et al, 2001). A patient (aged 58 years) with a mild clinical course and rare episodes of acute exacerbation was admitted to our division after a severe haemolytic crisis. He had a clinical history of mild anaemia since childhood, with no history of neonatal jaundice. In adulthood he had two episodes of haemolytic ‘crisis’ with severe anaemia (Hb 60–70 g/l), probably after exposure to oxidant stress. These episodes required blood transfusions. Familiar anamnesis was negative. During the years prior to the event the patient had been working as a hairdresser, using dyes containing naphthol, for about 30 years. We observed the patient after the second haemolytic crisis. Hb level was 85 g/l, and he had asthenia, pallor and mild icterus. No splenomegaly was present. Dermatitis was present on both hands, with deep cutaneous excoriations. Hypothesizing a possible association between the patient’s job and the haemolytic crises due to prolonged use of hair dyes, we advised the patient to use protective gloves and mask during hair dye preparation and recommended a reduction in working hours. No other haemolytic crises were observed. Subsequent blood counts over following months have shown a chronic haemolytic anaemia with haemoglobin levels between 100 and 110 g/l. Laboratory investigation revealed normal mean cell volume and mean cell haemoglobin concentration and elevated absolute reticulocyte counts ranging from 10% to 14%. No spherocytes were detected on peripheral blood smear review. G6PD enzyme level was reduced to 1Æ5 units/g haemoglobin (normal range 4Æ6–13Æ5). Testing for beta-thalassemia by sequencing and paroxysmal nocturnal haemoglobinuria clone were negative. Bone marrow aspiration showed only an erythroid hyperplasia. DNA sequencing of G6PD showed a novel mutation involving nucleotides 769 and 770 of exon 7, specifically C769A and G770T (Fig 1), designated G6PD Palermo. This transversion caused an amino acid substitution of arginine for methionine (R257M) in codon 257, which may change the protein structure, causing chronic haemolytic anaemia. After a few months the patient showed severe neutropenia with neutrophil counts <0Æ5 · 10/l. The chemical composition of the hair dyes used by the patient were found to contain benzene and toluene, both toxic for bone marrow. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzyme defect. The highest frequencies of G6PD deficiency are detected in Africa, Asia, Mediterranean region and in the Middle East. The striking similarity between the areas where G6PD deficiency is common and Plasmodium falciparum malaria is endemic provides circumstantial evidence that G6PD deficiency confers resistance against malaria (Ruwende & Hill, 1998; Cappellini & Fiorelli, 2008). In particular, malaria is endemic in Sicily and a previous study showed a rate of about 1Æ6% of G6PD-deficient males (56/3347), distributed throughout the island (Schiliro’ et al, 1979). We have previously described the occurrence of the mutations in the Sicilian population [G6PD Seattle (G6PD G844C), 37%; G6PD Mediterranean (G6PD C563T), 32%; G6PD*A(G6PD G202A/A376G), 18%; G6PD Santa Maria (G6PD A376G/A542T), 9%] (Sammarco P. (2008) unpublished observation). In some patients, variants of G6PD deficiency cause chronic haemolysis, leading to so-called congenital non-spherocytic haemolytic anaemia. The patient with G6PD Palermo R257M showed a congenital non-spherocytic anaemia caused by a Correspondence

A large view of CYP21 locus among Sicilians and other populations: identification of a novel CYP21A2 variant in Sicily

Background: Several mutations in CYP21 locus cause 21-hydroxylase deficiency (21-OHD). The most common mutations are widespread among different geographic areas and their frequencies have been also reported to differ among certain populations. Aim: To obtain a large view on the frequencies of the most common mutations in the CYP21 locus, in Sicily, in the Mediterranean and other major geographic areas worldwide. Subjects and methods: Three hundred and eight unrelated CYP21A2 alleles leading 21-OHD in Sicily were genetically typed and compared with other series previously reported in Sicily and in surrounding regions. An analysis of the frequencies of the different geographic areas was also carried out. CYP21A2 typing was carried out using PCR-restriction fragment length polymorphism (RFLP), for the detection of the CYP21A2 deletion, while sequencing analysis was performed to evaluate all the missense/non-sense mutations. Results: Our study revealed that p.V281L (44.4%), 12splice (21.6%) and p.P30L (11.2%) were very frequent alleles, del8bp (0.4%) was found very rarely in Sicily and a novel mutation leading to non-classical phenotype, p.L198F, was also discovered in this population. Allele frequencies were found to be significantly different from previously observed frequencies in Sicily. In addition, here we present the most significant frequency modifications among different geographic areas worldwide. Conclusions: As the distribution of the disease CYP21A2 alleles is heterogeneous around the world, the knowledge of the relative distributions allows a better management of 21-OHD for fetuses and newborns in different geographic areas.