Claudia Hemmelmann

Statistical analysis of rare sequence variants: an overview of collapsing methods.

With the advent of novel sequencing technologies, interest in the identification of rare variants that influence common traits has increased rapidly. Standard statistical methods, such as the Cochrane‐Armitage trend test or logistic regression, fail in this setting for the analysis of unrelated subjects because of the rareness of the variants. Recently, various alternative approaches have been proposed that circumvent the rareness problem by collapsing rare variants in a defined genetic region or sets of regions. We provide an overview of these collapsing methods for association analysis and discuss the use of permutation approaches for significance testing of the data‐adaptive methods. Genet. Epidemiol. 35:S12–S17, 2011.

Anti-JC virus antibodies in a large German natalizumab-treated multiple sclerosis cohort

Objective: To investigate the rate of seropositivity of anti-JC virus (JCV) antibodies in a German multiple sclerosis (MS) cohort treated with natalizumab in the postmarketing setting and to assess anti-JCV serostatus in samples obtained before diagnosis of progressive multifocal leukoencephalopathy (PML). Methods: This was a blinded, retrospective cross-sectional and longitudinal analysis for anti-JCV antibodies using a confirmatory 2-step ELISA on 2,782 blood samples obtained from 2,253 patients nationwide for routine testing for anti-natalizumab antibodies during open-label treatment between 2007 and 2010. Results: Of the natalizumab-treated patients with MS, 58.8% tested positive for anti-JCV antibodies. The rate of seropositivity was higher in males and increased with age, with a plateau between age intervals 20–29 and 30–39 years. In longitudinal analyses, 19 of 194 (9.8%) patients converted from anti-JCV antibody-negative to seropositive status over 7.7 months; 4.7% reverted from antibody-positive to seronegative status over 7.9 months. Antibody levels, especially in the latter group, were low, indicating fluctuations around the lower cut point of the assay. Neither anti-JCV serostatus nor antibody levels were associated with immunosuppressive pretreatment, duration of natalizumab treatment, or anti-natalizumab antibodies. All samples obtained from 10 patients who developed PML were seropositive (13 samples before PML diagnosis [2.0–37.6 months]; 2 samples at diagnosis). Antibody levels in these samples were higher than those in samples from seropositive patients who did not develop PML. Conclusions: These data argue for the potential clinical utility of JCV serology for PML risk stratification. However, further investigations of fluctuations in serostatus and of antibody levels for a more precise understanding of the predictive value are warranted.

Mutant Parkin Impairs Mitochondrial Function and Morphology in Human Fibroblasts

Background Mutations in Parkin are the most common cause of autosomal recessive Parkinson disease (PD). The mitochondrially localized E3 ubiquitin-protein ligase Parkin has been reported to be involved in respiratory chain function and mitochondrial dynamics. More recent publications also described a link between Parkin and mitophagy. Methodology/Principal Findings In this study, we investigated the impact of Parkin mutations on mitochondrial function and morphology in a human cellular model. Fibroblasts were obtained from three members of an Italian PD family with two mutations in Parkin (homozygous c.1072delT, homozygous delEx7, compound-heterozygous c.1072delT/delEx7), as well as from two relatives without mutations. Furthermore, three unrelated compound-heterozygous patients (delEx3-4/duplEx7-12, delEx4/c.924C>T and delEx1/c.924C>T) and three unrelated age-matched controls were included. Fibroblasts were cultured under basal or paraquat-induced oxidative stress conditions. ATP synthesis rates and cellular levels were detected luminometrically. Activities of complexes I-IV and citrate synthase were measured spectrophotometrically in mitochondrial preparations or cell lysates. The mitochondrial membrane potential was measured with 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide. Oxidative stress levels were investigated with the OxyBlot technique. The mitochondrial network was investigated immunocytochemically and the degree of branching was determined with image processing methods. We observed a decrease in the production and overall concentration of ATP coinciding with increased mitochondrial mass in Parkin-mutant fibroblasts. After an oxidative insult, the membrane potential decreased in patient cells but not in controls. We further determined higher levels of oxidized proteins in the mutants both under basal and stress conditions. The degree of mitochondrial network branching was comparable in mutants and controls under basal conditions and decreased to a similar extent under paraquat-induced stress. Conclusions Our results indicate that Parkin mutations cause abnormal mitochondrial function and morphology in non-neuronal human cells.

Observation and execution of upper-limb movements as a tool for rehabilitation of motor deficits in paretic stroke patients: protocol of a randomized clinical trial

BackgroundEvidence exist that motor observation activates the same cortical motor areas that are involved in the performance of the observed actions. The so called “mirror neuron system” has been proposed to be responsible for this phenomenon. We employ this neural system and its capability to re-enact stored motor representations as a tool for rehabilitating motor control. In our new neurorehabilitative schema (videotherapy) we combine observation of daily actions with concomitant physical training of the observed actions focusing on the upper limbs. Following a pilot study in chronic patients in an ambulatory setting, we currently designed a new multicenter clinical study dedicated to patients in the sub-acute state after stroke using a home-based self-induced training. Within our protocol we assess 1) the capability of action observation to elicit rehabilitational effects in the motor system, and 2) the capacity of this schema to be performed by patients without assistance from a physiotherapist. The results of this study would be of high health and economical relevance.Methods/designA controlled, randomized, multicenter, paralleled, 6 month follow-up study will be conducted on three groups of patients: one group will be given the experimental treatment whereas the other two will participate in control treatments. All patients will undergo their usual rehabilitative treatment beside participation in the study. The experimental condition consists in the observation and immediate imitation of common daily hand and arm actions. The two parallel control groups are a placebo group and a group receiving usual rehabilitation without any trial-related treatment. Trial randomization is provided via external data management. The primary efficacy endpoint is the improvement of the experimental group in a standardized motor function test (Wolf Motor Function Test) relative to control groups. Further assessments refer to subjective and qualitative rehabilitational scores. This study has been reviewed and approved by the ethics committee of Aachen University.DiscussionThis therapy provides an extension of therapeutic procedures for recovery after stroke and emphasizes the importance of action perception in neurorehabilitation The results of the study could become implemented into the wide physiotherapeutic practice, for example as an ad on and individualized therapy.

Incidence of therapy-related acute leukaemia in mitoxantrone-treated multiple sclerosis patients in Germany

Background: The incidence of therapy-related acute leukaemia (TRAL) in mitoxantrone treatment in multiple sclerosis (MS) is controversially discussed. Methods and results: In a retrospective meta-analysis from six centres, we observed six cases of acute myeloid leukaemia (AML) (incidence 0.41% for patients with mean follow up after end of treatment of 3.6 years, n = 1.156; incidence 0.25% for all patients, n = 2.261). Potential influencing factors such as myelotoxic or glucocorticosteroid pretreatment/cotreatment were present in all but one case of TRAL. Between 1990 and 2010, 11 cases of TRAL were reported to the Drug Commission of the German Medical Association (estimated risk of 0.09–0.13%). Conclusions: Regional differences in reported TRAL incidence may point to confounding cofactors such as administration protocols and cotreatments.

New concepts of multiple tests and their use for evaluating high-dimensional EEG data

Recently, new concepts of type I error control in multiple comparisons have been proposed, in addition to FWE and FDR control. We introduce these criteria and investigate in simulations how the powers of corresponding test procedures for multiple endpoints depend on various quantities such as number and correlation of endpoints, percentage of false hypotheses, etc. We applied the different multiple tests to EEG coherence data. We compared the memory encoding of subsequently recalled and not recalled nouns. The results show that subsequently recalled nouns elicited significantly higher coherence than not recalled ones.

Genome-wide association study in musician's dystonia: A risk variant at the arylsulfatase G locus?

Musicians dystonia (MD) affects 1% to 2% of professional musicians and frequently terminates performance careers. It is characterized by loss of voluntary motor control when playing the instrument. Little is known about genetic risk factors, although MD or writers dystonia (WD) occurs in relatives of 20% of MD patients. We conducted a 2‐stage genome‐wide association study in whites. Genotypes at 557,620 single‐nucleotide polymorphisms (SNPs) passed stringent quality control for 127 patients and 984 controls. Ten SNPs revealed P < 10−5 and entered the replication phase including 116 MD patients and 125 healthy musicians. A genome‐wide significant SNP (P < 5 × 10−8) was also genotyped in 208 German or Dutch WD patients, 1,969 Caucasian, Spanish, and Japanese patients with other forms of focal or segmental dystonia as well as in 2,233 ethnically matched controls. Genome‐wide significance with MD was observed for an intronic variant in the arylsulfatase G (ARSG) gene (rs11655081; P = 3.95 × 10−9; odds ratio [OR], 4.33; 95% confidence interval [CI], 2.66‐7.05). rs11655081 was also associated with WD (P = 2.78 × 10−2) but not with any other focal or segmental dystonia. The allele frequency of rs11655081 varies substantially between different populations. The population stratification in our sample was modest (λ = 1.07), but the effect size may be overestimated. Using a small but homogenous patient sample, we provide data for a possible association of ARSG with MD. The variant may also contribute to the risk of WD, a form of dystonia that is often found in relatives of MD patients.

Epidemic Microclusters of Blood-Culture Proven Sepsis in Very-Low-Birth Weight Infants: Experience of the German Neonatal Network

Introduction We evaluated blood culture-proven sepsis episodes occurring in microclusters in very-low-birth-weight infants born in the German Neonatal Network (GNN) during 2009–2010. Methods Thirty-seven centers participated in GNN; 23 centers enrolled ≥50 VLBW infants in the study period. Data quality was approved by on-site monitoring. Microclusters of sepsis were defined as occurrence of at least two blood-culture proven sepsis events in different patients of one center within 3 months with the same bacterial species. For microcluster analysis, we selected sepsis episodes with typically cross-transmitted bacteria of high clinical significance including gram-negative rods and Enterococcus spp. Results In our cohort, 12/2110 (0.6%) infants were documented with an early-onset sepsis and 235 late-onset sepsis episodes (≥72 h of age) occurred in 203/2110 (9.6%) VLBW infants. In 182/235 (77.4%) late-onset sepsis episodes gram-positive bacteria were documented, while coagulase negative staphylococci were found to be the most predominant pathogens (48.5%, 95%CI: 42.01–55.01). Candida spp. and gram-negative bacilli caused 10/235 (4.3%, 95%CI: 1.68% –6.83%) and 43/235 (18.5%) late-onset sepsis episodes, respectively. Eleven microclusters of blood-culture proven sepsis were detected in 7 hospitals involving a total 26 infants. 16/26 cluster patients suffered from Klebsiella spp. sepsis. The median time interval between the first patient’s Klebsiella spp. sepsis and cluster cases was 14.1 days (interquartile range: 1–27 days). First patients in the cluster, their linked cases and sporadic sepsis events did not show significant differences in short term outcome parameters. Discussion Microclusters of infection are an important phenomenon for late-onset sepsis. Most gram-negative cluster infections occur within 30 days after the first patient was diagnosed and Klebsiella spp. play a major role. It is essential to monitor epidemic microclusters of sepsis in surveillance networks to adapt clinical practice, inform policy and further improve quality of care.

Decreased EEG coherence between prefrontal electrodes: a correlate of high language proficiency?

To investigate the influence of proficiency level on the cortical organization of foreign language processing, two groups of German speaking students, differing only in their proficiency in English as a second language, were subjected to EEG coherence analysis during foreign and native language processing (news reports, alpha1 frequency band). In the group with minor experience with English, coherence increase was observed with all electrode combinations, with left hemisphere (LH) predominance. In the high proficiency group, coherence increase was limited to temporal electrodes over LH. In the latter group only, coherence between prefrontal electrodes was significantly lower during the language tasks than during the baseline task (silence, noisy screen). Both results were obtained with foreign as well as native language processing. We suggest that reduced EEG coherence in highly proficient foreign language speakers reflects a more efficient operating strategy not only for their second, but also for their native language.

Multivariate tests for the evaluation of high-dimensional EEG data.

In this paper several multivariate tests are presented, in particular permutation tests, which can be used in multiple endpoint problems as for example in comparisons of high-dimensional vectors of EEG data. We have investigated the power of these tests using artificial data in simulations and real EEG data. It is obvious that no one multivariate test is uniformly most powerful. The power of the different methods depends in different ways on the correlation between the endpoints, on the number of endpoints for which differences exist and on other factors. Based on our findings, we have derived rules of thumb regarding under which configurations a particular test should be used. In order to demonstrate the properties of different multivariate tests we applied them to EEG coherence data. As an example for the paired samples case, we compared the 171-dimensional coherence vectors observed for the alpha1-band while processing either concrete or abstract nouns and obtained significant global differences for some sections of time. As an example for the unpaired samples case, we compared the coherence vectors observed for language students and non-language students who processed an English text and found a significant global difference.