A walk in the PARC: developing and implementing 21st century chemical risk assessment in Europe.

Current approaches for the assessment of environmental and human health risks due to exposure to chemical substances have served their purpose reasonably well. Nevertheless, the systems in place for different uses of chemicals are faced with various challenges, ranging from a growing number of chemicals to changes in the types of chemicals and materials produced. This has triggered global awareness of the need for a paradigm shift, which in turn has led to the publication of new concepts for chemical risk assessment and explorations of how to translate these concepts into pragmatic approaches. As a result, next-generation risk assessment (NGRA) is generally seen as the way forward. However, incorporating new scientific insights and innovative approaches into hazard and exposure assessments in such a way that regulatory needs are adequately met has appeared to be challenging. The European Partnership for the Assessment of Risks from Chemicals (PARC) has been designed to address various challenges associated with innovating chemical risk assessment. Its overall goal is to consolidate and strengthen the European research and innovation capacity for chemical risk assessment to protect human health and the environment. With around 200 participating organisations from all over Europe, including three European agencies, and a total budget of over 400 million euro, PARC is one of the largest projects of its kind. It has a duration of seven years and is coordinated by ANSES, the French Agency for Food, Environmental and Occupational Health & Safety.

Atomic force microscopy for quantitative understanding of peptide-induced lipid bilayer remodeling.

A number of peptides are known to bind lipid bilayer membranes and cause these natural barriers to leak in an uncontrolled manner. Though membrane permeabilizing peptides play critical roles in cellular activity and may have promising future applications in the therapeutic arena, significant questions remain about their mechanisms of action. The atomic force microscope (AFM) is a single molecule imaging tool capable of addressing lipid bilayers in near-native fluid conditions. The apparatus complements traditional assays by providing local topographic maps of bilayer remodeling induced by membrane permeabilizing peptides. The information garnered from the AFM includes direct visualization and statistical analyses of distinct bilayer remodeling modes such as highly localized pore-like voids in the bilayer and dispersed thinned membrane regions. Colocalization of distinct remodeling modes can be studied. Here we examine recent work in the field and outline methods used to achieve precise AFM image data. Experimental challenges and common pitfalls are discussed as well as techniques for unbiased analysis including the Hessian blob detection algorithm, bootstrapping, and the Bayesian information criterion. When coupled with robust statistical analyses, high precision AFM data is poised to advance understanding of an important family of peptides that cause poration of membrane bilayers.

DNAscan: personal computer compatible NGS analysis, annotation and visualisation.

BACKGROUND: Next Generation Sequencing (NGS) is a commonly used technology for studying the genetic basis of biological processes and it underpins the aspirations of precision medicine. However, there are significant challenges when dealing with NGS data. Firstly, a huge number of bioinformatics tools for a wide range of uses exist, therefore it is challenging to design an analysis pipeline. Secondly, NGS analysis is computationally intensive, requiring expensive infrastructure, and many medical and research centres do not have adequate high performance computing facilities and cloud computing is not always an option due to privacy and ownership issues. Finally, the interpretation of the results is not trivial and most available pipelines lack the utilities to favour this crucial step. RESULTS: We have therefore developed a fast and efficient bioinformatics pipeline that allows for the analysis of DNA sequencing data, while requiring little computational effort and memory usage. DNAscan can analyse a whole exome sequencing sample in 1 h and a 40x whole genome sequencing sample in 13 h, on a midrange computer. The pipeline can look for single nucleotide variants, small indels, structural variants, repeat expansions and viral genetic material (or any other organism). Its results are annotated using a customisable variety of databases and are available for an on-the-fly visualisation with a local deployment of the gene.iobio platform. DNAscan is implemented in Python. Its code and documentation are available on GitHub: https://github.com/KHP-Informatics/DNAscan . Instructions for an easy and fast deployment with Docker and Singularity are also provided on GitHub. CONCLUSIONS: DNAscan is an extremely fast and computationally efficient pipeline for analysis, visualization and interpretation of NGS data. It is designed to provide a powerful and easy-to-use tool for applications in biomedical research and diagnostic medicine, at minimal computational cost. Its comprehensive approach will maximise the potential audience of users, bringing such analyses within the reach of non-specialist laboratories, and those from centres with limited funding available.

Rare variants analysis of cutaneous malignant melanoma genes in Parkinson's disease.

A shared genetic susceptibility between cutaneous malignant melanoma (CMM) and Parkinson's disease (PD) has been suggested. We investigated this by assessing the contribution of rare variants in genes involved in CMM to PD risk. We studied rare variation across 29 CMM risk genes using high-quality genotype data in 6875 PD cases and 6065 controls and sought to replicate findings using whole-exome sequencing data from a second independent cohort totaling 1255 PD cases and 473 controls. No statistically significant enrichment of rare variants across all genes, per gene, or for any individual variant was detected in either cohort. There were nonsignificant trends toward different carrier frequencies between PD cases and controls, under different inheritance models, in the following CMM risk genes: BAP1, DCC, ERBB4, KIT, MAPK2, MITF, PTEN, and TP53. The very rare TYR p.V275F variant, which is a pathogenic allele for recessive albinism, was more common in PD cases than controls in 3 independent cohorts. Tyrosinase, encoded by TYR, is the rate-limiting enzyme for the production of neuromelanin, and has a role in the production of dopamine. These results suggest a possible role for another gene in the dopamine-biosynthetic pathway in susceptibility to neurodegenerative Parkinsonism, but further studies in larger PD cohorts are needed to accurately determine the role of these genes/variants in disease pathogenesis.

Body site matters: an evaluation and application of a novel histological methodology on the quantification of mucous cells in the skin of Atlantic salmon, Salmo salar L.

Mucous cell size and distribution were investigated in the skin of five salmon using a novel stereology-based methodology: one (48 cm) fish to test 15 tissue treatment combinations on measures of cell area and density on the dorsolateral region and, using the most suitable treatment, we mapped mucous cell differences between body regions on four (52 cm) salmon, comprising a male and a female on each of two diets. The section site, decalcification, embedding medium and plane of sectioning all impacted significantly on mucous cell size, whereas mucous cell density is more robust. There were highly significant differences in both mucosal density and mean mucous cell size depending on body site: the dorsolateral skin of the four salmon had significantly denser (about 8% of skin area) and larger (mean about 160 µm(2)) mucous cells, whereas the lowest mean density (about 4%) and smallest mean area (115 µm(2)) were found on the head. We found that 100 random measurements may be sufficient to distinguish differences >7 µm(2) in mean mucous cell areas. The results further suggest that salmon exhibit a dynamic repeatable pattern of mucous cell development influenced by sex, diet and possibly strain and season.

The 14q22.2 colorectal cancer variant rs4444235 shows cis-acting regulation of BMP4.

Common genetic variation at human 14q22.2 tagged by rs4444235 is significantly associated with colorectal cancer (CRC) risk. Re-sequencing was used to comprehensively annotate the 17kb region of strong linkage disequilibrium encompassing rs4444235. Through bioinformatic analyses using H3K4Me1, H3K4Me3, and DNase-I hypersensitivity chromatin signatures and evolutionary conservation we identified seven candidate disease-causing single-nucleotide polymorphisms mapping to six regions within the 17-kb region predicted to have regulatory potential. Reporter gene studies of these regions demonstrated that the element to which rs4444235 maps acts as an allele-specific transcriptional enhancer. Allele-specific expression studies in CRC cell lines heterozygous for rs4444235 showed significantly increased expression of bone morphogenetic protein-4 (BMP4) associated with the risk allele (P<0.001). These data provide evidence for a functional basis for the non-coding risk variant rs4444235 at 14q22.2 and emphasizes the importance of genetic variation in the BMP pathway genes as determinants of CRC risk.

CASP8 variants D302H and -652 6N ins/del do not influence the risk of colorectal cancer in the United Kingdom population.

Polymorphisms in CASP8 at 2q33.1 have been associated with the risk of developing cancer, specifically, the D302H variant (rs1045485) with breast cancer in the European population and the -652 6N ins/del promoter variant (rs3834129) with multiple tumours including colorectal cancer (CRC) in the Chinese population. We evaluated the relationship between -652 6N ins/del and D302H variants and risk of developing CRC in the UK population by genotyping 4016 cases and 3749 controls. Both variants showed no evidence of an association with risk of developing CRC (P=0.42 and 0.22, respectively). In contrast, the recently identified CRC susceptibility allele rs6983267 mapping to 8q24 was significantly associated with disease risk (P=8.94 x 10(-8)). It is thus very unlikely that variation in CASP8 defined by -652 6N ins/del or D302H influences the risk of CRC in European populations. The implications of our findings both in terms of population-specific effects and publication bias are discussed.

Rapid word-learning in normal-hearing and hearing-impaired children: effects of age, receptive vocabulary, and high-frequency amplification.

OBJECTIVE: This study examined rapid word-learning in 5- to 14-year-old children with normal and impaired hearing. The effects of age and receptive vocabulary were examined as well as those of high-frequency amplification. Novel words were low-pass filtered at 4 kHz (typical of current amplification devices) and at 9 kHz. It was hypothesized that (1) the children with normal hearing would learn more words than the children with hearing loss, (2) word-learning would increase with age and receptive vocabulary for both groups, and (3) both groups would benefit from a broader frequency bandwidth. DESIGN: Sixty children with normal hearing and 37 children with moderate sensorineural hearing losses participated in this study. Each child viewed a 4-minute animated slideshow containing 8 nonsense words created using the 24 English consonant phonemes (3 consonants per word). Each word was repeated 3 times. Half of the 8 words were low-pass filtered at 4 kHz and half were filtered at 9 kHz. After viewing the story twice, each child was asked to identify the words from among pictures in the slide show. Before testing, a measure of current receptive vocabulary was obtained using the Peabody Picture Vocabulary Test (PPVT-III). RESULTS: The PPVT-III scores of the hearing-impaired children were consistently poorer than those of the normal-hearing children across the age range tested. A similar pattern of results was observed for word-learning in that the performance of the hearing-impaired children was significantly poorer than that of the normal-hearing children. Further analysis of the PPVT and word-learning scores suggested that although word-learning was reduced in the hearing-impaired children, their performance was consistent with their receptive vocabularies. Additionally, no correlation was found between overall performance and the age of identification, age of amplification, or years of amplification in the children with hearing loss. Results also revealed a small increase in performance for both groups in the extended bandwidth condition but the difference was not significant at the traditional p = 0.05 level. CONCLUSIONS: The ability to learn words rapidly appears to be poorer in children with hearing loss over a wide range of ages. These results coincide with the consistently poorer receptive vocabularies for these children. Neither the word-learning or receptive-vocabulary measures were related to the amplification histories of these children. Finally, providing an extended high-frequency bandwidth did not significantly improve rapid word-learning for either group with these stimuli.

Evidence suggesting that Homo neanderthalensis contributed the H2 MAPT haplotype to Homo sapiens.

The tau (MAPT) locus exists as two distinct clades, H1 and H2. The H1 clade has a normal linkage disequilibrium structure and is the only haplotype found in all populations except those derived from Caucasians. The H2 haplotype is the minor haplotype in Caucasian populations and is not found in other populations. It shows no recombination over a region of 2 Mb with the more common H1 haplotype. The distribution of the haplotype and analysis of the slippage of dinucleotide repeat markers within the haplotype suggest that it entered Homo sapiens populations between approx. 10000 and 30000 years ago. However, sequence comparison of the H2 haplotype with the H1 haplotype and with the chimp sequence suggests that the common founder of the H1 and H2 haplotypes was far earlier than this. We suggest that the H2 haplotype is derived from Homo neanderthalensis and entered H. sapiens populations during the co-existence of these species in Europe from approx. 45000 to 18000 years ago and that the H2 haplotype has been under selection pressure since that time, possibly because of the role of this H1 haplotype in neurodegenerative disease.

The H1c haplotype at the MAPT locus is associated with Alzheimer's disease.

Although it is clear that microtubule associated protein tau (MAPT) is involved in Alzheimer's disease (AD) pathology, it has not been clear whether it is involved genetically. We have recently examined the MAPT locus in progressive supranuclear palsy and found that a haplotype (H1c) on the background of the well-described H1 clade is associated with PSP. Here we report that the same haplotype is associated with the risk of AD in two autopsy confirmed series of cases with ages at death >65 years.

Linkage disequilibrium fine mapping and haplotype association analysis of the tau gene in progressive supranuclear palsy and corticobasal degeneration.

BACKGROUND: The haplotype H1 of the tau gene, MAPT, is highly associated with progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). OBJECTIVE: To investigate the pathogenic basis of this association. METHODS: Detailed linkage disequilibrium and common haplotype structure of MAPT were examined in 27 CEPH trios using validated HapMap genotype data for 24 single nucleotide polymorphisms (SNPs) spanning MAPT. RESULTS: Multiple variants of the H1 haplotype were resolved, reflecting a far greater diversity of MAPT than can be explained by the H1 and H2 clades alone. Based on this, six haplotype tagging SNPs (htSNPs) that capture 95% of the common haplotype diversity were used to genotype well characterised PSP and CBD case-control cohorts. In addition to strong association with PSP and CBD of individual SNPs, two common haplotypes derived from these htSNPs were identified that are highly associated with PSP: the sole H2 derived haplotype was underrepresented and one of the common H1 derived haplotypes was highly associated, with a similar trend observed in CBD. There were powerful and highly significant associations with PSP and CBD of haplotypes formed by three H1 specific SNPs. This made it possible to define a candidate region of at least approximately 56 kb, spanning sequences from upstream of MAPT exon 1 to intron 9. On the H1 haplotype background, these could harbour the pathogenic variants. CONCLUSIONS: The findings support the pathological evidence that underlying variations in MAPT could contribute to disease pathogenesis by subtle effects on gene expression and/or splicing. They also form the basis for the investigation of the possible genetic role of MAPT in Parkinson's disease and other tauopathies, including Alzheimer's disease.

Strong association of the Saitohin gene Q7 variant with progressive supranuclear palsy.

Recent reports are inconclusive in showing that the Q7R polymorphism of the novel Saitohin gene, nested in intron 9 of the tau gene, is associated with AD. The authors show that this polymorphism is in complete linkage disequilibrium with the extended tau H1/H2 haplotype and that the Q variant and QQ genotype of Q7R are strongly associated with progressive supranuclear palsy, implicating it as a possibly important pathogenic candidate.

Hearing loss in children and adults: audiometric configuration, asymmetry, and progression.

OBJECTIVE: The purpose of this study was to characterize the sensorineural hearing losses of a group of children and adults along three parameters important to the hearing instrument fitting process: 1) audiometric configuration, 2) asymmetry of loss between ears, and 3) progression of loss over several years. DESIGN: Audiograms for 248 60- and 61-yr-old adults and 227 6-yr-old children were obtained from the audiological database at Boys Town National Research Hospital. Based on right-ear air-conduction thresholds, the configurations were assigned to one of six categories: sloping, rising, flat, u-shaped, tent-shaped, and other. Left- and right-ear thresholds were compared to determine asymmetry of loss. Progression of loss was evaluated for 132 children for whom additional audiograms over an 8-yr period were available. RESULTS: In general, the children's hearing losses were more evenly distributed across configuration categories while most of the adult's audiograms were sloping or u-shaped in configuration. The variability of loss at each frequency was greater for the children than the adults for all configuration categories. Asymmetrical losses were more common and the degree of asymmetry at each frequency was more extensive among the children than the adults. A small number of children showed either improved or deteriorated hearing levels over time. In those children for whom progressive hearing loss occurred, no frequency was more vulnerable than another. CONCLUSIONS: The results of the present study suggest that substantial differences in audiological characteristics exist between children and adults. Implications for amplification include the development of appropriate fitting protocols for unusual audiometric configurations as well as protocols for binaural amplification in cases of asymmetric hearing losses.

Effect of stimulus bandwidth on the perception of /s/ in normal- and hearing-impaired children and adults.

Recent studies with adults have suggested that amplification at 4 kHz and above fails to improve speech recognition and may even degrade performance when high-frequency thresholds exceed 50-60 dB HL. This study examined the extent to which high frequencies can provide useful information for fricative perception for normal-hearing and hearing-impaired children and adults. Eighty subjects (20 per group) participated. Nonsense syllables containing the phonemes /s/, /f/, and /O/, produced by a male, female, and child talker, were low-pass filtered at 2, 3, 4, 5, 6, and 9 kHz. Frequency shaping was provided for the hearing-impaired subjects only. Results revealed significant differences in recognition between the four groups of subjects. Specifically, both groups of children performed more poorly than their adult counterparts at similar bandwidths. Likewise, both hearing-impaired groups performed more poorly than their normal-hearing counterparts. In addition, significant talker effects for /s/ were observed. For the male talker, optimum performance was reached at a bandwidth of approximately 4-5 kHz, whereas optimum performance for the female and child talkers did not occur until a bandwidth of 9 kHz.

Recognition of speech produced in noise.

A two-part study examined recognition of speech produced in quiet and in noise by normal hearing adults. In Part I 5 women produced 50 sentences consisting of an ambiguous carrier phrase followed by a unique target word. These sentences were spoken in three environments: quiet, wide band noise (WBN), and meaningful multi-talker babble (MMB). The WBN and MMB competitors were presented through insert earphones at 80 dB SPL. For each talker, the mean vocal level, long-term average speech spectra, and mean word duration were calculated for the 50 target words produced in each speaking environment. Compared to quiet, the vocal levels produced in WBN and MMB increased an average of 14.5 dB. The increase in vocal level was characterized by increased spectral energy in the high frequencies. Word duration also increased an average of 77 ms in WBN and MMB relative to the quiet condition. In Part II, the sentences produced by one of the 5 talkers were presented to 30 adults in the presence of multi-talker babble under two conditions. Recognition was evaluated for each condition. In the first condition, the sentences produced in quiet and in noise were presented at equal signal-to-noise ratios (SNR(E)). This served to remove the vocal level differences between the speech samples. In the second condition, the vocal level differences were preserved (SNR(P)). For the SNR(E) condition, recognition of the speech produced in WBN and MMB was on average 15% higher than that for the speech produced in quiet. For the SNR(P) condition, recognition increased an average of 69% for these same speech samples relative to speech produced in quiet. In general, correlational analyses failed to show a direct relation between the acoustic properties measured in Part I and the recognition measures in Part II.

Child maltreatment: risk of adjustment problems and dating violence in adolescence.

OBJECTIVE: To examine the relationship between child maltreatment, clinically relevant adjustment problems, and dating violence in a community sample of adolescents. METHOD: Adolescents from 10 high schools (N= 1,419; response rate = 62%) in southwestern Ontario completed questionnaires that assessed past maltreatment, current adjustment, and dating violence. Logistic regression was used to compare maltreated and nonmaltreated youths across outcome domains. RESULTS: One third (n = 462) of the school sample reported levels of maltreatment above the cutoff score on the Childhood Trauma Questionnaire. Girls with a history of maltreatment had a higher risk of emotional distress compared with girls without such histories (e.g., odds ratios [OR] for anger, depression, anxiety, and posttraumatic stress-related problems were 7.1, 7.2, 9.3, and 9.8, respectively). They were also at greater risk of violent and nonviolent delinquency (OR = 2.7) and carrying concealed weapons (OR = 7.1). Boys with histories of maltreatment were 2.5 to 3.5 times as likely to report clinical levels of depression, posttraumatic stress, and overt dissociation as were boys without a maltreatment history. They also had a significantly greater risk of using threatening behaviors (OR = 2.8) or physical abuse (OR = 3.4) against their dating partners. CONCLUSIONS: Maltreatment is a significant risk factor for adolescent maladjustment and shows a differential pattern for male and female adolescents.

Childhood maltreatment, posttraumatic stress symptomatology, and adolescent dating violence: considering the value of adolescent perceptions of abuse and a trauma mediational model.

The present study, utilizing both a child protective services and high school sample of midadolescents, examined the issue of self-report of maltreatment as it relates to issues of external validity (i.e., concordance with social worker ratings). reliability (i.e.. overlap with an alternate child maltreatment self-report inventory; association of a self-labeling item as "abused" with their subscale item counterparts), and construct validity (i.e., the association of maltreatment with posttraumatic stress symptomatology and dating violence). Relevant theoretical work in attachment, trauma, and relationship violence points to a mediational model, whereby the relationship between childhood maltreatment and adolescent dating violence would be expected to be accounted for by posttraumatic stress symptomatology. In the high school sample, 1329 adolescents and, in the CPS sample, 224 youth on the active caseloads completed comparable questionnaires in the three domains of interest. For females only, results supported a mediational model in the prediction of dating violence in both samples. For males, child maltreatment and trauma symptomatology added unique contributions to predicting dating violence. with no consistent pattern emerging across samples. When considering the issue of self-labeling as abused. CPS females who self-labeled had higher posttraumatic stress symptomatology and dating violence victimization scores than did their nonlabeling, maltreated counterparts for emotional maltreatment. These results point to the need for ongoing work in understanding the process of disclosure and how maltreatment experiences are consciously conceptualized.

Brain-derived neurotrophic factor differentially regulates excitatory and inhibitory synaptic transmission in hippocampal cultures.

Brain-derived neurotrophic factor (BDNF) has been postulated to be a key signaling molecule in regulating synaptic strength and overall circuit activity. In this context, we have found that BDNF dramatically increases the frequency of spontaneously initiated action potentials in hippocampal neurons in dissociated culture. Using analysis of unitary synaptic transmission and immunocytochemical methods, we determined that chronic treatment with BDNF potentiates both excitatory and inhibitory transmission, but that it does so via different mechanisms. BDNF strengthens excitation primarily by augmenting the amplitude of AMPA receptor-mediated miniature EPSCs (mEPSCs) but enhances inhibition by increasing the frequency of mIPSC and increasing the size of GABAergic synaptic terminals. In contrast to observations in other systems, BDNF-mediated increases in AMPA-receptor mediated mEPSC amplitudes did not require activity, because blocking action potentials with tetrodotoxin for the entire duration of BDNF treatment had no effect on the magnitude of this enhancement. These forms of synaptic regulations appear to be a selective action of BDNF because intrinsic excitability, synapse number, and neuronal survival are not affected in these cultures. Thus, although BDNF induces a net increase in overall circuit activity, this results from potentiation of both excitatory and inhibitory synaptic drive through distinct and selective physiological mechanisms.

The relation between stimulus context, speech audibility, and perception for normal-hearing and hearing-impaired children.

In this study, the influence of stimulus context and audibility on sentence recognition was assessed in 60 normal-hearing children, 23 hearing-impaired children, and 20 normal-hearing adults. Performance-intensity (PI) functions were obtained for 60 semantically correct and 60 semantically anomalous sentences. For each participant, an audibility index (AI) was calculated at each presentation level, and a logistic function was fitted to rau-transformed percent-correct values to estimate the SPL and AI required to achieve 70% performance. For both types of sentences, there was a systematic age-related shift in the PI functions, suggesting that young children require a higher AI to achieve performance equivalent to that of adults. Improvement in performance with the addition of semantic context was statistically significant only for the normal-hearing 5-year-olds and adults. Data from the hearing-impaired children showed age-related trends that were similar to those of the normal-hearing children, with the majority of individual data falling within the 5th and 95th percentile of normal. The implications of these findings in terms of hearing-aid fitting strategies for young children are discussed.