R. Cerone

Minor facial anomalies in combined methylmalonic aciduria and homocystinuria due to a defect in cobalamin metabolism.

Methylmalonic acidaemia and homocystinuria (MMA/HC) is a very rare inborn error of cellular metabolism of cobalamin (Cbl), that results in functional defects of both methylmalonyl-CoA mutase and methionine synthase because of deficient synthesis of 5-deoxyadenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl). AdoCbl is the cofactor of methylmalonyl-CoA mutase, the enzyme catalysing the intramitochondrial isomerization of methylmalonyl-CoA to succinyl-CoA, while MeCbl is the cofactor of the cytosolic remethylation from homocysteine to methionine with recycling of the folate derivative tetrahydrofolate (THF). Seven different defects of the intracellular metabolism of Cbl have been described; of these seven, three forms involve the synthesis of both AdoCbl and MeCbl and are distinct genetically and biochemically, namely Cbl-C (the most frequent variant), Cbl-D and Cbl-F. Age of onset and clinical findings vary widely among the three complementation groups and among known patients with Cbl-C disease: the affected patients generally show feeding difficulties, failure to thrive, neurological dysfunction (developmental delay, hypotonia, seizures, extrapyramidal signs in childhood, dementia, psychosis), haematological changes, ocular abnormalities and renal involvement. The acute neonatal onset is severe and can lead to death (Fenton and Rosenblatt 1995; Rosenblatt et al 1997)). To date, minor facial anomalies have been reported only in two cases of the Cbl-F form, but not in the other variants of combined MMA and HC, whereas peculiar facial dysmorphism has been described in several different metabolic diseases such as peroxisomal disorders, carbohydrate glycoprotein deficiency syndrome, Smith-Lemli-Opitz syndrome, mevalonic aciduria, pyruvate dehydrogenase deficiency and respiratory chain deficiencies. We report here our personal observation of these clinical findings in seven patients with MMA and HC.

Pregnancy and tyrosinaemia type II

A female patient with tyrosinaemia type II is reported having undergone two untreated pregnancies. During pregnancies, plasma tyrosine was raised. The outcomes of both offspring show that maternal tyrosinaemia may have an adverse effect on the developing fetus.

Fumarate hydratase deficiency

Fumarate hydratase deÐciency E. Bonioli1*, A. Di Stefano1, V . Peri1, U. Caruso1, R. Cerone1, E. L amantea3, F. Taroni3 and C. Bellini2 1 Istituto di Clinica Pediatrica, 2 Istituto di Puericultura e Medicina Neonatale, University of Genoa ; 3 Laboratorio di Patologia Cellulare, Dipartimento di Biochimica e Genetica, Istituto Neurologico Besta, Milan, Italy * Correspondence : Istituto di Clinica Pediatrica dellÏUniversità, Largo G. Gaslini, 5, 16147 Genova, Italy

A spectrum of LMX1B mutations in Nail-Patella syndrome: New point mutations, deletion, and evidence of mosaicism in unaffected parents

Purpose: Nail-Patella syndrome (MIM 161200) is a rare autosomal dominant disorder characterized by hypoplastic or absent patellae, dystrophic nails, dysplasia of the elbows, and iliac horn. In 40% of cases, a glomerular defect is present and, less frequently, ocular damage is observed. Inter- and intrafamilial variable expressivity of the clinical phenotype is a common finding. Mutations in the human LMX1B gene have been demonstrated to be responsible for Nail-Patella syndrome in around 80% of cases.Methods: Standard polymerase chain reaction and sequencing methods were used for mutation and single nucleotide polymorphism identification and control of cloned sequences. Array-CGH (Agilent, 244A Kit) was used for detection of deletions. Standard cloning techniques and the Snapshot method were used for analysis of mosaicism.Results: In this study, we present the results of LMX1B screening of 20 Nail-Patella syndrome patients. The molecular defect was found in 17 patients. We report five novel mutations and a ∼2 Mb deletion in chromosome 9q encompassing the entire LMX1B gene in a patient with a complex phenotype. We present evidence of somatic mosaicism in unaffected parents in two cases, which, to our knowledge, are the first reported cases of inheritance of a mutated LMX1B allele in Nail-Patella syndrome patients from a mosaic parent.Conclusion: The study of the described case series provides some original observations in an “old” genetic disorder.

Testing for tetrahydrobiopterin responsiveness in patients with hyperphenylalaninemia due to phenylalanine hydroxylase deficiency.

IntroductionPharmacological levels of the phenylalanine hydroxylase enzyme cofactor, tetrahydrobiopterin (BH4), reduce plasma phenylalanine levels in some patients with phenylketonuria (PKU), providing the first pharmacological therapy for PKU. Responsiveness to this therapy must be determined empirically through a BH4 loading test or trial. The authors have analyzed the loading tests currently in use in light of the numerous factors that can influence their results. Sapropterin dihydrochloride is a stable, synthetic form of BH4 approved for treatment of PKU in responsive patients.MethodsAn expert panel identified evidence from published reports of clinical experience. Reports of research involving at least 25 patients and published in English were considered.ResultsIn all, 14 studies met both criteria; eight employing the sapropterin dihydrochloride preparation from Schircks Laboratories and six the sapropterin dihydrochloride preparation from Biomarin/Merck Serono.ConclusionThe arbitrary responsiveness definition of a >30% reduction in blood phenylalanine appears to be a good compromise between sensitivity and specificity for the initial screening test. However, individual patient characteristics should be considered when interpreting results, especially in patients with low baseline phenylalanine levels.

Why do premature newborn infants display elevated blood adenosine levels

Our preliminary data show high levels of adenosine in the blood of very low birth weight (VLBW) infants, positively correlating to their prematurity (i.e. body weight class). This prompted us to look for a mechanism promoting such impressive adenosine increase. We hypothesized a correlation with oxygen challenge. In fact, it is recognized that either oxygen lack or its excess contribute to the pathogenesis of the injuries of prematurity, such as retinopathy (ROP) and periventricular white matter lesions (PWMI). The optimal concentration of oxygen for resuscitation of VLBW infants is currently under revision. We propose that the elevated adenosine blood concentrations of VLBW infants recognizes two sources. The first could be its activity-dependent release from unmyelinated brain axons. Adenosine in this respect would be an end-product of the hypometabolic VLBW newborn unmyelinated axon intensely firing in response to the environmental stimuli consequent to premature birth. Adenosine would be eventually found in the blood due to blood-brain barrier immaturity. In fact, adenosine is the primary activity-dependent signal promoting differentiation of premyelinating oligodendrocyte progenitor cells (OPC) into myelinating cells in the Central Nervous System, while inhibiting their proliferation and inhibiting synaptic function. The second, would be the ecto-cellular ATP synthesized by the endothelial cell plasmalemma exposed to ambient oxygen concentrations due to premature breathing, especially in lung. ATP would be rapidly transformed into adenosine by the ectonucleotidase activities such as NTPDase I (CD39), and NT5E (CD73). An ectopic extra-mitochondrial aerobic ATP synthetic ability was reported in many cell plasma-membranes, among which endothelial cells. The potential implications of the cited hypotheses for the neonatology area would be great. The amount of oxygen administration for reviving of newborns would find a molecular basis for its assessment. VLBW infants may be regarded as those in which premature exposure to ambient oxygen concentrations and oxidative stress causes a premature functioning of the extra-mitochondrial oxidative phosphorylation primarily in axons and endothelium. Adenosine may become a biomarker of prematurity risk, whose implications further studies may assess.

Common criteria among States for storage and use of dried blood spot specimens after newborn screening

Biological samples collected in biobanks are a resource with significant research potential. The Italian Joint Group CNB - CNBBSV (National Committee of Bioethics - National Committee for Biosecurity, Biotechnologies and Life Sciences) published a document reporting recommendations on storage and use of dried blood spot (DBS) and on the development of a National Network of Regional Newborn Screening Repositories for collection of residual DBS. Several ethical questions (about consent, possible use of genetic information, unanticipated possible usages for research purposes) rise from residual newborn screening specimens collections. Moreover, legal and ethical controversies are accentuated by the conflicts between the interests of sample donors, biobank holders, researchers and the public. To overcome these difficulties the identification of a few criteria for storage and research usage of DBS is crucial.

von willebrand factor multimer composition is modified following oral methionine load in women with thrombosis, but not in healthy women

Hyperhomocysteinemia is associated with an increased risk of venous and arterial thrombosis, probably by inducing endothelial damage. Von Willebrand factor (VWF) is an endothelial marker protein. It is a plasma multimeric molecule that plays a thrombophilic role. Our purpose was to investigate VWF changes in patients with thrombosis following oral methionine load. We evaluated homocysteine levels and VWF parameters (plasma levels, activity, proteolysis fragments, and multimer composition) before and after methionine load in 42 women with venous or arterial thrombosis and in 36 healthy women. Methionine load induced mild hyperhomocysteinemia in 10 patients and two controls. No changes in VWF levels and activity were observed, but an increased amount of VWF proteolysis fragments was found post-load in patients and controls. VWF multimer composition was unaffected in controls, while a decrease of the largest VWF multimers was found in women with thrombosis. Homocysteine levels inversely correlated with the amount of the largest multimers in hyperhomocysteinemic patients. Large VWF molecules were probably released from endothelial cells following load, and rapidly cleaved by the specific VWF-cleaving protease. VWF proteolysis was enhanced in mild hyperhomocysteinemic patients, thus leading to downregulation of VWF size to smaller multimers.

Isolated sulphite oxidase deficiency: clinical and biochemical features in an Italian patient

Summary: A patient with isolated sulphite oxidase deficiency presented with seizures at 12 h of life and followed a severe course, dying at 10 months of age. There was mild facial dysmorphism and the brain showed multiple cystic lesions.

Genotype–phenotype correlation in dihydropteridine reductase deficiency

Inherited deficiency of dihydropteridine reductase (DHPR, EC 1.66.99.7) impairs the regeneration of tetrahydrobiopterin (BH 4 ), the essential cofactor of phenylalanine (Phe) (PAH, EC 1.14.16.1), tyrosine (Tyr) (TYH, EC 1.14.16.2) and tryptophan (Trp) (TRH, EC 1.14.16.4) hydroxylases, which is oxidized to qBH 2 during a coupled reaction with these enzymes. The main metabolic derangements caused by DHPR deficiency (McKusick 261630) are hyperphenylalaninaemia and impaired production of monoamine neurotransmitters derived from Tyr and Trp dopamine, noradrenaline and serotonin. Untreated patients can early develop a severe and progressive neurological picture (Blau et al 1996a). The control of hyperphenylalaninaemia and biogenic amine deficiency is necessary to improve their prognosis, together with folinic acid supplementation to avoid folate depletion (Spada et al 1996). However, in some DHPR patients a milder phenotype has been described characterized by absent neurological signs. These patients respond to a BH 4 monotherapy or do not require any treatment (Blau et al 1992). The DHPR gene (QDPR), on 4p15.3, includes seven exons (Dianzani et al 1998). It encodes for a protein of 244 amino acids, active as a homodimer. So far, 21 mutations have been described in QDPR uniformly scattered throughout the coding region (Dianzani et al 1998; Smooker et al 1999), with different mutations having different effects on the protein, as determined by in vitro studies (Smooker et al 1993; Zhang et al 1996). Most mutations have been found in single chromosomes. So far, only five of them have been identified more than once (de Sanctis et al 1996). In the present study we evaluated genotype-phenotype correlation in 21 completely characterized DHPR patients. Molecular, biochemical and clinical data on DHPR-deficient patients are stored in the BIOMDB and BIODEF database (Blau et al 1996b).