Genome‐wide association study identifies loci associated with liability to alcohol and drug dependence that is associated with variability in reward‐related ventral striatum activity in African‐ and European‐Americans

Genetic influences on alcohol and drug dependence partially overlap, however, specific loci underlying this overlap remain unclear. We conducted a genome‐wide association study (GWAS) of a phenotype representing alcohol or illicit drug dependence (ANYDEP) among 7291 European‐Americans (EA; 2927 cases) and 3132 African‐Americans (AA: 1315 cases) participating in the family‐based Collaborative Study on the Genetics of Alcoholism. ANYDEP was heritable (h ² in EA = 0.60, AA = 0.37). The AA GWAS identified three regions with genome‐wide significant (GWS; P < 5E‐08) single nucleotide polymorphisms (SNPs) on chromosomes 3 (rs34066662, rs58801820) and 13 (rs75168521, rs78886294), and an insertion‐deletion on chromosome 5 (chr5:141988181). No polymorphisms reached GWS in the EA. One GWS region (chromosome 1: rs1890881) emerged from a trans‐ancestral meta‐analysis (EA + AA) of ANYDEP, and was attributable to alcohol dependence in both samples. Four genes (AA: CRKL, DZIP3, SBK3; EA: P2RX6) and four sets of genes were significantly enriched within biological pathways for hemostasis and signal transduction. GWS signals did not replicate in two independent samples but there was weak evidence for association between rs1890881 and alcohol intake in the UK Biobank. Among 118 AA and 481 EA individuals from the Duke Neurogenetics Study, rs75168521 and rs1890881 genotypes were associated with variability in reward‐related ventral striatum activation. This study identified novel loci for substance dependence and provides preliminary evidence that these variants are also associated with individual differences in neural reward reactivity. Gene discovery efforts in non‐European samples with distinct patterns of substance use may lead to the identification of novel ancestry‐specific genetic markers of risk.     

Arrhythmogenic right ventricular dysplasia/cardiomyopathy associated with mutations in the desmosomal gene desmocollin-2

Arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C) is an inherited myocardial disorder associated with arrhythmias, heart failure, and sudden death. To date, mutations in four genes encoding major desmosomal proteins (plakoglobin, desmoplakin, plakophilin-2, and desmoglein-2) have been implicated in the pathogenesis of ARVD/C. We screened 77 probands with ARVD/C for mutations in desmocollin-2 (DSC2), a gene coding for a desmosomal cadherin. Two heterozygous mutations--a deletion and an insertion--were identified in four probands. Both mutations result in frameshifts and premature truncation of the desmocollin-2 protein. For the first time, we have identified mutations in desmocollin-2 in patients with ARVD/C, a finding that is consistent with the hypothesis that ARVD/C is a disease of the desmosome.     

Improved fermentation processes for NS0 cell lines expressing human antibodies and glutamine synthetase

To meet the increasing requirement for therapeutic antibodies to conduct clinical trials, an enhanced culture medium and fed-batch process was developed for GS-NS0 cell lines. This process was shown to produce high concentrations of monoclonal antibodies for several cell lines expressing different antibodies. Cells were adapted to growth in a glutamine- and serum-free medium containing bovine serum albumin (BSA), cholesterol, and transferrin. A number of amino acids were found to be depleted during cell culture. The concentrations of these amino acids were increased, and further cell culture analyses were performed. This process of cell growth and analysis was repeated over multiple cycles until no depletion was detected. This resulted in an amino acid supplement that was shown to be generic and enhanced antibody productivity up to 5-fold for the three cell lines tested. Transferrin was replaced using tropolone, a lipophilic iron chelator and ferric ammonium citrate. Cell growth was equivalent to that in transferrin-containing medium over the wide ranges tested. A concentrated feed solution, based on the amino acid supplement and the components of the serum- and protein-free supplements, was formulated. Addition of this feed in response to metabolic requirements resulted in a harvest titer a further 2-fold higher than the enhanced culture medium. Harvest antibody titers of up to 600 mg/L were achieved for three cell lines expressing different antibodies, representing an increase of 10-fold over the starting concentrations.     

Phase-resetting curve determines how BK currents affect neuronal firing

BK channels are large conductance potassium channels gated by calcium and voltage. Paradoxically, blocking these channels has been shown experimentally to increase or decrease the firing rate of neurons, depending on the neural subtype and brain region. The mechanism for how this current can alter the firing rates of different neurons remains poorly understood. Using phase-resetting curve (PRC) theory, we determine when BK channels increase or decrease the firing rates in neural models. The addition of BK currents always decreases the firing rate when the PRC has only a positive region. When the PRC has a negative region (type II), BK currents can increase the firing rate. The influence of BK channels on firing rate in the presence of other conductances, such as I _m and I _h , as well as with different amplitudes of depolarizing input, were also investigated. These results provide a formal explanation for the apparently contradictory effects of BK channel antagonists on firing rates.     

A New Statistic to Evaluate Imputation Reliability

Background

As the amount of data from genome wide association studies grows dramatically, many interesting scientific questions require imputation to combine or expand datasets. However, there are two situations for which imputation has been problematic: (1) polymorphisms with low minor allele frequency (MAF), and (2) datasets where subjects are genotyped on different platforms. Traditional measures of imputation cannot effectively address these problems.

Methodology/Principal Findings

We introduce a new statistic, the imputation quality score (IQS). In order to differentiate between well-imputed and poorly-imputed single nucleotide polymorphisms (SNPs), IQS adjusts the concordance between imputed and genotyped SNPs for chance. We first evaluated IQS in relation to minor allele frequency. Using a sample of subjects genotyped on the Illumina 1 M array, we extracted those SNPs that were also on the Illumina 550 K array and imputed them to the full set of the 1 M SNPs. As expected, the average IQS value drops dramatically with a decrease in minor allele frequency, indicating that IQS appropriately adjusts for minor allele frequency. We then evaluated whether IQS can filter poorly-imputed SNPs in situations where cases and controls are genotyped on different platforms. Randomly dividing the data into “cases” and “controls”, we extracted the Illumina 550 K SNPs from the cases and imputed the remaining Illumina 1 M SNPs. The initial Q-Q plot for the test of association between cases and controls was grossly distorted (λ = 1.15) and had 4016 false positives, reflecting imputation error. After filtering out SNPs with IQS<0.9, the Q-Q plot was acceptable and there were no longer false positives. We then evaluated the robustness of IQS computed independently on the two halves of the data. In both European Americans and African Americans the correlation was >0.99 demonstrating that a database of IQS values from common imputations could be used as an effective filter to combine data genotyped on different platforms.

Conclusions/Significance

IQS effectively differentiates well-imputed and poorly-imputed SNPs. It is particularly useful for SNPs with low minor allele frequency and when datasets are genotyped on different platforms.     

NADPH oxidase-derived reactive oxygen species in cardiac pathophysiology

Chronic heart failure, secondary to left ventricular hypertrophy or myocardial infarction, is a condition with increasing morbidity and mortality. Although the mechanisms underlying the development and progression of this condition remain a subject of intense interest, there is now growing evidence that redox-sensitive pathways play an important role. This article focuses on the involvement of reactive oxygen species derived from a family of superoxide-generating enzymes, termed NADPH oxidases (NOXs), in the pathophysiology of ventricular hypertrophy, the accompanying interstitial fibrosis and subsequent heart failure. In particular, the apparent ability of the different NADPH oxidase isoforms to define the response of a cell to a range of physiological and pathophysiological stimuli is reviewed. If confirmed, these data would suggest that independently targeting different members of the NOX family may hold the potential for therapeutic intervention in the treatment of cardiac disease.