Rare copy number variation in cerebral palsy

Recent studies have established the role of rare copy number variants (CNVs) in several neurological disorders but the contribution of rare CNVs to cerebral palsy (CP) is not known. Fifty Caucasian families having children with CP were studied using two microarray designs. Potentially pathogenic, rare (<1% population frequency) CNVs were identified, and their frequency determined, by comparing the CNVs found in cases with 8329 adult controls with no known neurological disorders. Ten of the 50 cases (20%) had rare CNVs of potential relevance to CP; there were a total of 14 CNVs, which were observed in <0.1% (<8/8329) of the control population. Eight inherited from an unaffected mother: a 751-kb deletion including FSCB, a 1.5-Mb duplication of 7q21.13, a 534-kb duplication of 15q11.2, a 446-kb duplication including CTNND2, a 219-kb duplication including MCPH1, a 169-kb duplication of 22q13.33, a 64-kb duplication of MC2R, and a 135-bp exonic deletion of SLC06A1. Three inherited from an unaffected father: a 386-kb deletion of 12p12.2-p12.1, a 234-kb duplication of 10q26.13, and a 4-kb exonic deletion of COPS3. The inheritance was unknown for three CNVs: a 157-bp exonic deletion of ACOX1, a 693-kb duplication of 17q25.3, and a 265-kb duplication of DAAM1. This is the first systematic study of CNVs in CP, and although it did not identify de novo mutations, has shown inherited, rare CNVs involving potentially pathogenic genes and pathways requiring further investigation.     

Menopausal hormone therapy in Germany: Results of three national surveys from 1997 to 2003

Objective

To determine regional variations in the prevalence and applied therapy regimes of current menopausal hormone therapy (HT) in Germany.

Methods

Three population-based surveys, analysing data of 45–74 years old women, were compared: The Study of Health in Pomerania (SHIP; 1123 participants; northeast Germany, October 1997–May 2001), Cooperative Health Research in the Augsburg Region Survey 2000 (KORA; 1253 participants; south Germany, October 1999–April 2001) and Heinz Nixdorf Recall Study (HNR; 2257 participants; west Germany, December 2000–August 2003). A standardized interview technique provided data on current medication.

Results

The age-standardized prevalence of HT was 17.0% (95% confidence interval (CI): 14.9–19.1) in SHIP, 25.9% (95% CI: 23.6–28.3) in KORA and 24.7% (95% CI: 22.9–26.4) in HNR. Mean average time of intake of HT was 5.1 (SHIP), 7.5 (KORA) and 10.1 years (HNR). The use of estrogen plus progestogen combinations was equally common in all three surveys with proportions of about 15%, the use of unopposed estrogen in KORA and HNR was twice as high as in SHIP. In all three surveys oral estradiol was taken most often. Transdermal estradiol was preferred by KORA women whereas conjugated estrogens were taken most frequently by HNR women.

Conclusions

Compared to northeast Germany HT was more often applied in the south and west of Germany. HT as long-term therapy was more common in West than in East Germany. In each study region there was a specific pattern of used HT components.     

Long-term neurological outcome of term-born children treated with two or more anti-epileptic drugs during the neonatal period

Background

Neonatal seizures may persist despite treatment with multiple anti-epileptic drugs (AEDs).

Objective

To determine in term-born infants with seizures that required two or more AEDs, whether treatment efficacy and/or the underlying disorder were related to neurological outcome.

Design/methods

We included 82 children (born 1998-2006) treated for neonatal seizures. We recorded mortality, aetiology of seizures, the number of AEDs required, achievement of seizure control, and amplitude-integrated-EEG (aEEG) background patterns. Follow-up consisted of an age-adequate neurological examination. Surviving children were classified as normal, having mild neurological abnormalities, or cerebral palsy (CP).

Results

Forty-seven infants (57%) had status epilepticus. The number of AEDs was not related to neurological outcome. Treatment with three or four AEDs as opposed to two showed a trend towards an increased risk of a poor outcome, i.e., death or CP, odds ratio (OR) 2.74; 95% confidence interval (CI) 0.98-7.69; P = .055. Failure to achieve seizure control increased the risk of poor outcome, OR 6.77; 95%-CI 1.42-32.82, P = .016. Persistently severely abnormal aEEG background patterns also increased this risk, OR 3.19; 95%-CI 1.90-5.36; P < .001. In a multivariate model including abnormal aEEG background patterns, failure to achieve seizure control nearly reached significance towards an increased risk of poor outcome, OR 5.72, 95%-CI 0.99-32.97, P = .051. We found no association between seizure aetiology and outcome.

Conclusions

In term-born infants with seizures that required two or more AEDs outcome was poorer if seizure control failed. The number of AEDs required to reach seizure control and seizure aetiology had limited prognostic value.     

A comprehensive survey for polymorphisms in the bovine IFN-gamma gene reveals a highly polymorphic intronic DNA sequence allowing improved genotyping of Bovinae.

This study aimed to identify interferon-gamma (IFN-gamma) gene variants in cattle for diagnostic purposes. Therefore, the entire bovine IFN-gamma gene (BoIFNG) and 2605 bp of its promoter DNA were sequenced. The BoIFNG DNA sequence conforms to the published part of Bo-IFN-gamma cDNA. Primer extension experiments show the presence of a 5' extension of exon 1 by 42 nucleotides (nt). One SINE element (Bov-A2) is located in the 5'-region, and two SINE elements (Bov-tA, Bov-B) are contained in the 3'-region of BoIFNG. The variants were detected by comparative sequence analysis of PCR amplicons from different bovine species. Four polymorphic mononucleotide repeats are situated in the promoter and in intron 1. Four distinct series of single nucleotide polymorphisms (SNP) were found in functionally important regions of BoIFNG. The region between the two intron 1 microsatellites contains the highest density of SNPs in Bos taurus breeds. One G-T transversion in the coding region of exon 1 causes a Gly(14) to Val(14) exchange in the BoIFNG signal peptide of different bovine species. A G-A transition in exon 2 encodes a Ser(19) to Asn(19) change in the mature protein of the Tibetan yak. Genotyping of randomly sampled Holstein Friesian cows at selected SNPs and of both intron 1 microsatellites revealed two dominant BoIFNG microhaplotypes. The detected SNPs improve the recently reported genotyping system of cattle.     

Clinical and molecular basis of classical lissencephaly: Mutations in the LIS1 gene (PAFAH1B1)

Classical lissencephaly (LIS) and subcortical band heterotopia (SBH) are related cortical malformations secondary to abnormal migration of neurons during early brain development. Approximately 60% of patients with classical LIS, and one patient with atypical SBH have been found to have deletions or mutations of the LIS1 gene, located on 17p13.3. This gene encodes the LIS1 or PAFAH1B1 protein with a coiled‐coil domain at the N‐terminus and seven WD40 repeats at the C‐terminus. It is highly conserved between species and has been shown to interact with multiple proteins involved with cytoskeletal dynamics, playing a role in both cellular division and motility, as well as the regulation of brain levels of platelet activating factor. Here we report 65 large deletions of the LIS1 gene detected by FISH and 41 intragenic mutations, including four not previously reported, the majority of which have been found as a consequence of the investigation of 220 children with LIS or SBH by our group. All intragenic mutations are de novo, and there have been no familial recurrences. Eight‐eight percent (36/41) of the mutations result in a truncated or internally deleted protein—with missense mutations found in only 12% (5/41) thus far. Mutations occurred throughout the gene except for exon 7, with clustering of three of the five missense mutations in exon 6. Only five intragenic mutations were recurrent. In general, the most severe LIS phenotype was seen in patients with large deletions of 17p13.3, with milder phenotypes seen with intragenic mutations. Of these, the mildest phenotypes were seen in patients with missense mutations. Hum Mutat 19:4–15, 2002. © 2001 Wiley‐Liss, Inc.     

Norepinephrine transporter (NET) promoter and 5'-UTR polymorphisms: association analysis in panic disorder

Several biochemical and pharmacological studies suggest that the catecholaminergic system involving the norepinephrine transporter (NET) is relevant for the pathogenesis of panic disorder. Three single nucleotide polymorphisms in the promoter or untranslated 5' region of the NET gene were investigated by means of RFLP analysis in a sample of 115 German patients with panic disorder and 115 matched controls. Statistical analysis failed to show association with the overall diagnosis of panic disorder. In the subgroup of patients with panic disorder without agoraphobia, however, two polymorphisms were found to be associated with the disease (G/C (rs2397771): p < 0.05; T/C (rs2242446): p < 0.01). While our data do not support a major function of the NET gene in the development of panic disorder, it may play a role in the subgroup of panic disorder without agoraphobia.     

Detection and calibration of microdeletions and microduplications by array-based comparative genomic hybridization and its applicability to clinical genetic testing.

PURPOSE: Genome-wide telomere screening by fluorescence in situ hybridization (FISH) has revealed that approximately 6% of unexplained mental retardation is due to submicroscopic telomere imbalances. However, the use of FISH for telomere screening is labor intensive and time consuming, given that 41 telomeres are interrogated. We have evaluated the use of array-based Comparative Genomic Hybridization (aCGH) as a more efficient tool for identifying telomere rearrangements. METHODS: In this study, 102 individuals with unexplained mental retardation, with either normal or abnormal FISH results, were selected for a blinded retrospective study using aCGH. Results between the two methodologies were compared to ascertain the ability of aCGH to be used in a clinical diagnostics setting. RESULTS: We detected 100% of all imbalances previously identified by FISH (n = 17) and identified two additional abnormalities, a 10q telomere duplication and an interstitial duplication of 22q11. Interphase FISH analysis verified all abnormal array results. We also demonstrated that aCGH can accurately calibrate the size of telomere imbalances by using an array with "molecular rulers" for the telomeric regions of 1p, 16p, 17p, and 22q. CONCLUSION: This study demonstrates that aCGH is an equivalent methodology to telomere FISH for detecting submicroscopic deletions. In addition, small duplications that are not easily visible by FISH can be accurately detected using aCGH. Because aCGH allows simultaneous interrogation of hundreds to thousands of DNA probes and is more amenable to automation, it offers an efficient and high-throughput alternative for detecting and calibrating unbalanced rearrangements, both of the telomere region, as well as other genomic locations.     

The t(14;18) in diffuse large B-cell lymphoma: correlation with germinal center-associated markers and clinical features.

The clinical and biologic relevance of the t(14;18) and features of germinal center (GC) differentiation in diffuse large B-cell lymphoma (DLBCL) remain controversial. The authors examined the association of t(14;18) with GC-associated markers and clinical features in 44 de novo DLBCLs (22 nodal and 22 primary extranodal). CD10, bcl-2, and bcl-6 were expressed in 50%, 62%, and 54% of cases respectively. There were no significant differences in expression of these markers between nodal and extranodal cases. Coexpression of CD10 and bcl-6 was seen in 12 of 41 cases, and was more frequent in nodal than extranodal DLBCL (9 of 21 vs. 3 of 20; P = 0.05). A CD10+/bcl-6+ phenotype was not significantly associated with bcl-2 expression, stage, complete remission rate, or survival. The t(14;18) was found in 7 of 44 (16%) cases (6 nodal, 1 extranodal; P = 0.09). It was associated with a CD10+/bcl-6+ phenotype (5 of 7 vs. 7 of 27; P = 0.015) and a trend toward more frequent bcl-6 expression (6 of 7 vs. 15 of 34; P = 0.09), but no association with bcl-2 expression, CD10, clinical stage, complete remission, or survival. Among nodal or high-stage (III-IV) DLBCL, cases with the t(14;18) showed a trend toward decreased survival (P = 0.12).     

Exercise rapidly increases eukaryotic elongation factor 2 phosphorylation in skeletal muscle of men

Protein synthesis in skeletal muscle is known to decrease during contractions but the underlying regulatory mechanisms are unknown. Here, the effect of exercise on skeletal muscle eukaryotic elongation factor 2 (eEF2) phosphorylation, a key component in protein translation machinery, was examined. Eight healthy men exercised on a cycle ergometer at a workload eliciting ∼67% peak pulmonary oxygen consumption     with skeletal muscle biopsies taken from the vastus lateralis muscle at rest as well as after 1, 10, 30, 60 and 90 min of exercise. In response to exercise, there was a rapid (i.e. < 1 min) 5- to 7-fold increase in eEF2 phosphorylation at Thr56 that was sustained for 90 min of continuous exercise. The in vitro activity of skeletal muscle eEF2 kinase was not altered by exercise indicating that the increased activity of eEF2 kinase to eEF2 is not mediated by covalent mechanisms. In support of this, the increase in AMPK activity was temporally unrelated to eEF2 phosphorylation. However, skeletal muscle eEF2 kinase was potently activated by Ca²⁺–calmodulin in vitro , suggesting that the higher eEF2 phosphorylation in working skeletal muscle is mediated by allosteric activation of eEF2 kinase by Ca²⁺ signalling via calmodulin. Given that eEF2 phosphorylation inhibits eEF2 activity and mRNA translation, these findings suggest that the inhibition of protein synthesis in contracting skeletal muscle is due to the Ca²⁺-induced stimulation of eEF2 kinase.     

The English version of the Verona medical interview classification system (VR-MICS). An assessment of its reliability and a comparative cross-cultural test of its validity

OBJECTIVE: This study aimed to assess the inter-rater and intra-rater reliability of the English translation of the original Italian version of the VR-MICS and to evaluate its sensitivity by comparing the coding of English and Italian general practice consultations with emotionally distressed and non-distressed patients, as defined by the 12-item General Health Questionnaire (GHQ-12). METHOD: Six male GPs from Manchester (UK) and six from Verona (Italy) each contributed five consultations, which were coded using the VR-MICS. Intra-rater and inter-rater reliability were assessed both for the division of interviews into speech units and the speech unit coding. Interaction and main effects of GHQ-12 status and nationality on patient and GP expressions were assessed by two-way ANOVA. RESULTS: Agreement indices for the division of speech units varied between 88-96 and 87-93% for GP and patient speech, respectively; those for coding categories between 88-91 and 82-86%, with Cohen's Kappa values between 0.86-0.91 and 0.80-0.85 for GP and patient speech, respectively. Cross-cultural comparisons of patient and GP speech showed no interaction effects between GHQ-12 status and nationality. The Italian GPs were more 'doctor-centred', while the UK GPs tended to use a more 'sharing' consulting style. Independent of nationality, distressed patients talked more, gave more psychosocial cues and increased amounts of positive talk compared to non-distressed patients. GPs in both settings, when interviewing distressed patients, reduced social conversation and increased psychosocial information-giving, checking questions and reassurance. CONCLUSION: The English translation of the VR-MICS showed satisfactory reliability indices and similar sensitivity to patients' verbal behaviours in relation to their emotional state in the two settings. PRACTICE IMPLICATIONS: The VR-MICS may be an useful coding instrument to support collaborative research on doctor-patient communication between the two countries.