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Brain tumours are the most common solid tumours in childhood. Half of these tumours occur in the posterior fossa, where surgical removal is complicated by the risk of cerebellar mutism syndrome, of which postoperative speech impairment (POSI) is a cardinal symptom, in up to 25% of patients. The surgical approach to midline tumours, mostly undertaken by transvermian or telovelar routes, has been proposed to influence the risk of POSI. We aimed to investigate the risk of developing POSI, the time course of its resolution, and its association with surgical approach and other clinical factors.

Usher syndrome (USH), the most prevalent cause of hereditary deafness–blindness, is an autosomal recessive and genetically heterogeneous disorder. Three clinical subtypes (USH1–3) are distinguishable based on the severity of the sensorineural hearing impairment, the presence or absence of vestibular dysfunction, and the age of onset of the retinitis pigmentosa. A total of 10 causal genes, 6 for USH1, 3 for USH2, and 1 for USH3, and an USH2 modifier gene, have been identified. A robust molecular diagnosis is required not only to improve genetic counseling, but also to advance gene therapy in USH patients. Here, we present an improved diagnostic strategy that is both cost- and time-effective. It relies on the sequential use of three different techniques to analyze selected genomic regions: targeted exome sequencing, comparative genome hybridization, and quantitative exon amplification. We screened a large cohort of 427 patients (139 USH1, 282 USH2, and six of undefined clinical subtype) from various European medical centers for mutations in all USH genes and the modifier gene. We identified a total of 421 different sequence variants predicted to be pathogenic, about half of which had not been previously reported. Remarkably, we detected large genomic rearrangements, most of which were novel and unique, in 9% of the patients. Thus, our strategy led to the identification of biallelic and monoallelic mutations in 92.7% and 5.8% of the USH patients, respectively. With an overall 98.5% mutation characterization rate, the diagnosis efficiency was substantially improved compared with previously reported methods.

Exercise has a potent insulin-sensitivity enhancing effect on skeletal muscle, but the intracellular mechanisms that mediate this effect are not well understood. In muscle, Ras-related C3 botulinum toxin substrate 1 (Rac1) regulates both insulin- and contraction-stimulated glucose transport and is dysregulated in insulin resistant muscle. However, whether Rac1 is involved in mediating enhanced insulin sensitivity after an acute bout of exercise is unresolved. To address this question, we investigated after exercise whole-body (insulin tolerance test) as well as muscle (insulin-stimulated 2-deoxyglucose transport in isolated soleus muscle) insulin sensitivity in inducible muscle-specific Rac1 knockout (mKO) and wild-type (WT) littermate mice. Previous exercise enhanced whole-body insulin sensitivity by 40% in WT mice and rescued the insulin intolerance in Rac1 mKO mice by improving whole-body insulin sensitivity by 230%. In agreement, previous exercise significantly improved insulin sensitivity by 20% in WT and by 40% in Rac1 mKO soleus muscles. These findings suggest that muscle Rac1 is dispensable for the insulin sensitizing effect of exercise. Moreover, insulin resistance in Rac1 mKO mice can be completely normalized by previous exercise explaining why insulin resistant patients can increase insulin action with exercise despite dysfunctional Rac1 activity in muscle.

Background:Exposure to secondhand smoke is associated with an increased risk of acute myocardial infarction (AMI). The positive impact of a smoking ban on AMI hospitalization rates has been demonstrated both inside and outside Europe. A national smoking ban (SB) was implemented in Denmark on 15 August 2007.Purpose:To evaluate the impact of the Danish SB on hospital admissions for AMI.Methods:Poisson regression models were used to analyse changes over time in AMI-admissions in Denmark. We investigated a seven year period: five years before and two years after implementation of the SB. We accounted for the variation in the population size and for seasonal trends. Potential confounders included were: gender, age and the incidence of type 2 diabetes (T2D).Results:A significant reduction in the number of AMI-admissions was found in the last three years of the study period after adjusting for the potential confounders. The significant reductions were found one year before the SB (relative rate (RR) = 0.86, 95% confidence interval (CI) 0.79–0.94), one year after the SB (RR = 0.77, 95% CI 0.71–0.85) and two years after the SB (RR = 0.77, 95% CI 0.70–0.84).Conclusions:A significant reduction in the number of AMI-admissions was found already one year before the SB after adjustment for the incidence of T2D. The results differ from most results found in similar studies throughout the world and may be explained by the incremental enactment of SBs in Denmark and the implementation of a nation-wide ban on industrially produced trans-fatty acids in food in 2004.

The Hutterian Bretheren is an isolated population living on the North American prairies, the current community exceeding 40,000 in number. Their unique genetic history has contributed to a founder effect, which is reflected in the Mendelian disorders present in this population today. Genetic studies in the Hutterite population have led to the identification of a number of genes over the last several years and highlights the power of this population for gene identification. However, for the more than 30 autosomal recessive conditions currently recognized in this population, the gene or Hutterite specific mutation remains to be identified for over half and novel autosomal recessive syndromes continue to be recognized. This review summarizes what is currently understood about the molecular etiology of the Mendelian disorders and highlights the cardinal features of those disorders that are unique to or over‐represented in this population. © 2008 Wiley‐Liss, Inc.

We identified a partial gene deletion of ATP7B in a patient with Wilson disease with hepatic onset. The deletion covered exon 20 including major parts of the flanking introns. The breakpoints were identified and the size of the deletion determined to be 2144 bp. The deletion is predicted to lead to a mutated protein product containing 45 aberrant amino acids after transmembrane domain 7, and lacking the transmembrane domain 8 as well as the entire C‐terminal cytoplasmic tail. This is the first time a partial gene deletion has been demonstrated in ATP7B. The patient presented at age 10 with hepatic manifestations, including severe jaundice, hepato‐splenomegaly, ascites, and spider naevi. The liver biopsy showed fibrosis and early signs of cirrhosis. There was a Kayser–Fleischer ring but no neurological manifestations. All symptoms disappeared with penicillamine therapy. This suggests that the C‐terminal cytoplasmatic tail of ATP7B, is not essential for its neurological function. Large deletions in ATP7B may be an overlooked cause of Wilson disease. Patients that are homozygotes for deletions may be valuable for the understanding of the function of various regions of the ATP7B protein. © 2005 Wiley‐Liss, Inc.

This work investigated a three-generation Menkes disease family, where germ-line mosaicism was suspected in the maternal grandmother of the index patient. She had given birth to 2 boys who died of suspected Menkes disease on the basis of clinical and photographic evidence. Biochemical analysis of the index patient confirmed the diagnosis of Menkes disease, and DNA analysis established a partial gene deletion (EX11_EX23del), involving exons 11–23 and the 3'-untranslated region (UTR) of ATP7A. A junction fragment was detectable by Southern blot analysis, which enabled carrier analysis. The mother was demonstrated to be a carrier, whereas analysis of lymphoblasts and skin fibroblasts from the maternal grandmother gave no indication of a partial gene deletion. No materials were available from the possibly affected maternal uncles. Further genetic analyses, including biochemical testing of the grandmother and haplotype analysis using four intragenic markers on DNA from selected members of the family, corroborated this finding. The combined results from DNA analyses showed that the grandmother had transmitted three different ATP7A haplotypes to her offspring: (1) the at-risk allele (CAB)-1 and the deletion; (2) the at-risk allele (CAB)-1 without deletion; and (3) the second allele (CAB)-2 without deletion. In conclusion, our study demonstrated segregation of Menkes disease within the family investigated that can best be explained by extensive germ-line mosaicism in the maternal grandmother. The finding of germ-line mosaicism has obvious implications for genetic counseling of Menkes disease families.

Purpose: Phenylketonuria (PKU) is an autosomal recessive disorder caused by mutations in the phenylalanine hydroxylase (PAH) gene. There have been more than 400 mutations identified in the PAH gene leading to variable degrees of deficiency in PAH activity, and consequently a wide spectrum of clinical severity. A pilot study was undertaken to examine the response to 6-R-l-erythro-5,6,7,8-tetrahydrobiopterin (BH4) in patients with atypical and classical PKU.Methods: PAH gene mutation analysis was performed using denaturing gradient gel electrophoresis and gene sequencing. Patients with classical, atypical, or mild PKU were orally given BH4 10 mg/kg. Blood phenylalanine and tyrosine levels were determined using tandem MS/MS at 0 hours, 4 hours, 8 hours, and 24 hours intervals.Results: Thirty-six patients were given a single oral dose of 10 mg/kg of BH4. Twenty one patients (58.33%) responded with a decrease in blood phenylalanine level. Of the patients that responded, 12 were classical, 7 atypical, and 2 mild. The mean decline in blood phenylalanine at 24 hours was > 30% of baseline. There were 15 patients who did not respond to the BH4challenge, 14 of those had classical and one had atypical PKU. Mapping the mutations that responded to BH4on the PAH enzyme showed that mutations were in the catalytic, regulatory, oligomerization, and BH4binding domains. Five patients responding to BH4had mutations not previously identified.Conclusion: The data presented suggest higher than anticipated number of PKU mutations respond to BH4, and such mutations are on all the domains of PAH.

Purpose: Phenylketonuria (PKU) is an autosomal recessive disorder caused by mutations in the phenylalanine hydroxylase (PAH) gene. There have been more than 400 mutations identified in the PAH gene leading to variable degrees of deficiency in PAH activity, and consequently a wide spectrum of clinical severity. A pilot study was undertaken to examine the response to 6-R-l-erythro-5,6,7,8-tetrahydrobiopterin (BH4) in patients with atypical and classical PKU.

In Escherichia coli there are two pathways for conversion of adenine into guanine nucleotides, both involving the intermediary formation of IMP. The major pathway involves conversion of adenine into hypoxanthine in three steps via adenosine and inosine, with subsequent phosphoribosylation of hypoxanthine to IMP. The minor pathway involves formation of ATP, which is converted via the histidine pathway to the purine intermediate 5-amino-4-imidazolecarboxamide ribonucleotide and, subsequently, to IMP. Here we describe E. coli mutants, in which a third pathway for conversion of adenine to IMP has been activated. This pathway was shown to involve direct deamination of adenine to hypoxanthine by a manganese-dependent adenine deaminase encoded by a cryptic gene, yicP, which we propose be renamed ade. Insertion elements, located from -145 to +13 bp relative to the transcription start site, activated the ade gene as did unlinked mutations in the hns gene, encoding the histone-like protein H-NS. Gene fusion analysis indicated that ade transcription is repressed more than 10-fold by H-NS and that a region of 231 bp including the ade promoter is sufficient for this regulation. The activating insertion elements essentially eliminated the H-NS-mediated silencing, and stimulated ade gene expression 2-3-fold independently of the H-NS protein.

Pyrimidine-requiring cddmutants of Escherichia colideficient in cytidine deaminase utilize cytidine as a pyrimidine source by an alternative pathway. This has been presumed to involve phosphorylation of cytidine to CMP by cytidine/uridine kinase and subsequent hydrolysis of CMP to cytosine and ribose 5-phosphate by a putative CMP hydrolase. Here we show that cytidine, in cddstrains, is converted directly to cytosine and ribose by a ribonucleoside hydrolase encoded by the previously uncharacterized gene ybeK, which we have renamed rihA. The RihA enzyme is homologous to the products of two unlinked genes, yeiKand yaaF, which have been renamed rihBand rihC, respectively. The RihB enzyme was shown to be a pyrimidine-specific ribonucleoside hydrolase like RihA, whereas RihC hydrolyzed both pyrimidine and purine ribonucleosides. The physiological function of the ribonucleoside hydrolases in wild-type E. colistrains is enigmatic, as their activities are paralleled by the phosphorolytic activities of the nucleoside phosphorylases, and a triple mutant lacking all three hydrolytic activities grew normally. Furthermore, enzyme assays and lacZgene fusion analysis indicated that rihBwas essentially silent unless activated by mutation, whereas rihAand rihCwere poorly expressed in glucose medium due to catabolite repression.

The ATP and GTP pools of Escherichia colihave recently been reported to increase approximately 10-fold with increasing growth rates in the range from 0.4 to 1.4 generations/hour (Gaal, T., Bartlett, M. S., Ross, W., Turnbough, C. L., and Gourse, R. L. (1997) Science278, 2092–2097). Moreover, it was proposed that this variation of the nucleotide pools, particularly the ATP pool, might be responsible for the well known growth rate-dependent regulation of rRNA synthesis in E. coli.To test this hypothesis we have measured the nucleoside triphosphate pools as a function of growth rate for several E. colistrains. We found that the size of all four RNA precursor pools are essentially invariant with growth rate, in the range from 0.5 to 2.3 generations/hour. Nevertheless we observed the expected growth rate-dependent increase of RNA accumulation in these strains. In light of these results, it seems unlikely that nucleotide pool variations should be responsible for the growth rate-dependent regulation of rRNA synthesis.

Molecular variation among major histocompatibility complex (MHC) class I (B-F) proteins from B-homozygous chickens is apparently caused by C-terminal variation. Analysis of the total B-F protein pool revealed substantial heterogeneity with two or three molecular mass constituents, each being comprised by several isoelectric focusing variants. This heterogeneity could not be reduced by enzymatic deglycosylation. By contrast, proteolytic removal of a small (Mr 1000–4000) fragment from the a chain resulted in the generation of a Mr 36 000 fragment, common to all the molecular mass variants. Unlike the parent proteins, the Mr 36 000 fragment derived from isolated variants yielded identical, simple patterns in two-dimensional gel electrophoresis and identical finger prints in peptide mapping. This, together with N-terminal amino acid sequencing, as well as comparison of hydrophobicity properties of fragments obtained by gradual proteolytic digestion, indicated that the small peptide responsible for the major B-F heterogeneity was situated in the intracellular, C-terminal part.

To examine the therapeutic consequences of diagnostk tests for AIDS-related infections, case records from 33 deceased AIDS patients were reviewed; 23 were autopsied. Determination of serum antibody titres was not important. In particular, there was no relation between titres and isolation of cytomegalovirus (CMV); isolation attempts or possibly antigen determination would be better. Samples for CMV isolation were obtained from 31 patients; ony 7 were negative. Blood, faeces, and particularly sputum cultures gave a low yield; the number of such examinations could be reduced considerably. However, 4/7 disseminated infections with atypical mycobacteria were only revealed at autopsy, despite numerous cultures in vivo. Liver biopsies were not helpful. Diagnostic procedures for Pneumocystis carinii pneumonia by lung biopsy caused pneumothorax in 3/15 patients; bronchoalveolar lavage or treatment/prophylaxis without diagnosis could be considered. In 8 autopsies, microscopy was suggestive of cerebral toxoplasmosis, but only 1 patient had presented important clinical symptoms. We suggest a schedule with regular microbiologic and parasitic examinations and few antibody tests, but with more antigen tests.

Two maternal half-brothers presented with huge cephalic hematoma, fatal in one. Skin morphology disclosed lack of elastic fibres. Their maternal uncle is moderately mentally handicapped and has extensive connective tissue disorders. In all these patients, an identical missense mutation in the ATP7Agene was found and confirmed Menkes' disease.