Initial experience on abdominal photon-counting computed tomography in clinical routine: general image quality and dose exposure.

OBJECTIVES: Photon-counting computed tomography has lately found its way into clinical routine. The new technique could offer substantial improvements regarding general image quality, image noise, and radiation dose reduction. This study evaluated the first abdominal examinations in clinical routine and compared the results to conventional computed tomography. METHODS: In this single-center retrospective study, 66 patients underwent photon-counting and conventional abdominal CT. Four radiologists assessed general image quality, image noise, and image artifacts. Signal-to-noise ratio and dose properties of both techniques within the clinical application were compared. An ex vivo phantom study revealed the radiobiological impact by means of DNA double-strand break foci in peripheral blood cells by enumerating γ-H2AX+53BP1 foci. RESULTS: General image quality in accordance with the Likert scale was found superior for photon-counting CT (4.74 ± 0.46 vs. 4.25 ± 0.54; p < 0.001). Signal-to-noise ratio (p < 0.001) and also dose exposure were higher for photon-counting CT (DLP: 419.2 ± 162.2 vs. 372.3 ± 236.6 mGy*cm; p = 0.0435). CT exposure resulted in significantly increased DNA damage in comparison to sham control (p < 0.001). Investigation of the average foci per cell and radiation-induced foci numbers revealed significantly elevated numbers (p = 0.004 and p < 0.0001, respectively) after photon-counting CT. CONCLUSION: Photon-counting CT in abdominal examinations showed superior results regarding general image quality and signal-to-noise ratio in clinical routine. However, this seems to be traded for a significantly higher dose exposure and corresponding double-strand break frequency. Optimization of standard protocols in further clinical applications is required to find a compromise regarding picture quality and dose exposure. KEY POINTS: • Photon-counting computed tomography promises to enhance the diagnostic potential of medical imaging in clinical routine. • Retrospective single-center study showed superior general image quality accompanied by higher dose exposure in initial abdominal PCCT protocols compared to state-of-the-art conventional CT. • A simultaneous ex vivo phantom study revealed correspondingly increased frequencies of DNA double-strand breaks after PCCT.

Gene expression for biodosimetry and effect prediction purposes: promises, pitfalls and future directions - key session ConRad 2021.

PURPOSE: In a nuclear or radiological event, an early diagnostic or prognostic tool is needed to distinguish unexposed from low- and highly exposed individuals with the latter requiring early and intensive medical care. Radiation-induced gene expression (GE) changes observed within hours and days after irradiation have shown potential to serve as biomarkers for either dose reconstruction (retrospective dosimetry) or the prediction of consecutively occurring acute or chronic health effects. The advantage of GE markers lies in their capability for early (1-3 days after irradiation), high-throughput, and point-of-care (POC) diagnosis required for the prediction of the acute radiation syndrome (ARS). CONCLUSIONS: As a key session of the ConRad conference in 2021, experts from different institutions were invited to provide state-of-the-art information on a range of topics including: (1) Biodosimetry: What are the current efforts to enhance the applicability of this method to perform retrospective biodosimetry? (2) Effect prediction: Can we apply radiation-induced GE changes for prediction of acute health effects as an approach, complementary to and integrating retrospective dose estimation? (3) High-throughput and point-of-care diagnostics: What are the current developments to make the GE approach applicable as a high-throughput as well as a POC diagnostic platform? (4) Low level radiation: What is the lowest dose range where GE can be used for biodosimetry purposes? (5) Methodological considerations: Different aspects of radiation-induced GE related to more detailed analysis of exons, transcripts and next-generation sequencing (NGS) were reported.

Impact of medical imaging on the epigenome - low-dose exposure in the course of computed tomography does not induce detectable changes of DNA-methylation profiles in peripheral blood cells.

BACKGROUND: Computed tomography (CT) is a main contributor to artificial low-dose exposure. Understanding the biological effects induced by CT exposure and their dependency on the characteristics of photon spectra is essential for knowledge-driven risk assessment. In a previous gene expression study, we have identified upregulation of AEN, BAX, DDB2, EDA2R and FDXR after ex vivo exposure with single-energy CT and dual-energy CT (DECT). In this study, we focused on CT-induced changes of DNA methylation. This epigenetic modification of DNA is a central regulator of gene expression and instrumental in preserving genome integrity. Previous studies reported focal hypermethylation and global hypomethylation after exposure with doses above 100 mSv, however, the effect of low dose exposure on DNA methylation is hardly explored. MATERIALS AND METHODS: DNA was isolated from peripheral blood of three healthy individuals 6 h after ex vivo exposition to single-energy (80 kV and 150 kV) and DECT (80 kV/Sn150 kV) with a calculated effective dose of 7.0 ± 0.08 mSv. The experimental setting was identical to the one used in our previous gene expression study enabling a direct comparison of gene expression results with changes of DNA methylation identified in this study. DNA methylation was analyzed by high-throughput sequencing of bisulfite-treated DNA targeted methylation sequencing. RESULTS: Unsupervised hierarchical clustering based on DNA methylation profiles of all samples created three distinct clusters. Formation of these three clusters was solely determined by the origin of samples, indicating the absence of prominent irradiation-associated changes of DNA methylation. In line with this observation, inter-individual comparison of non-irradiated samples revealed 1163, 1224 and 4550 significant differentially methylated regions (DMRs), respectively, whereas the pairwise comparison of irradiated and non-irradiated samples failed to identify irradiation-induced DMRs in any of the three probands. This even applied to the genomic regions harboring AEN, BAX, DDB2, EDA2R and FDXR, the five genes known to be upregulated by CT exposure. CONCLUSIONS: CT exposure with various photon spectra did not result in detectable changes of DNA methylation. However, minor effects in a subpopulation of irradiated cells cannot be ruled out. Thus, future studies with extended observation intervals are needed to investigate DNA methylation changes that are induced by indirect effects at later points of time or become detectable by clonal expansion of affected cells. Moreover, our data suggest that DNA methylation analysis is less sensitive in detecting immediate effects of low-dose irradiation when compared to gene expression analysis.

Gene expression changes and DNA damage after ex vivo exposure of peripheral blood cells to various CT photon spectra.

Dual-energy CT provides enhanced diagnostic power with similar or even reduced radiation dose as compared to single-energy CT. Its principle is based on the distinct physical properties of low and high energetic photons, which, however, may also affect the biological effectiveness and hence the extent of CT-induced cellular damage. Therefore, a comparative analysis of biological effectiveness of dual- and single-energy CT scans with focus on early gene regulation and frequency of radiation-induced DNA double strand breaks (DSBs) was performed. Blood samples from three healthy individuals were irradiated ex vivo with single-energy (80 kV and 150 kV) and dual-energy tube voltages (80 kV/Sn150kV) employing a modern dual source CT scanner resulting in Volume Computed Tomography Dose Index (CTDIvol) of 15.79-18.26 mGy and dose length product (DLP) of 606.7-613.8 mGy*cm. Non-irradiated samples served as a control. Differential gene expression in peripheral blood mononuclear cells was analyzed 6 h after irradiation using whole transcriptome sequencing. DSB frequency was studied by 53BP1 + γH2AX co-immunostaining and microscopic evaluation of their focal accumulation at DSBs. Neither the analysis of gene expression nor DSB frequency provided any evidence for significantly increased biological effectiveness of dual-energy CT in comparison to samples irradiated with particular single-energy CT spectra. Relative to control, irradiated samples were characterized by a significantly higher rate of DSBs (p < 0.001) and the shared upregulation of five genes, AEN, BAX, DDB2, FDXR and EDA2R, which have already been suggested as radiation-induced biomarkers in previous studies. Despite steadily decreasing doses, CT diagnostics remain a genotoxic stressor with impact on gene regulation and DNA integrity. However, no evidence was found that varying X-ray spectra of CT impact the extent of cellular damage.

Genomic Adaption and Mutational Patterns in a HaCaT Subline Resistant to Alkylating Agents and Ionizing Radiation.

Sulfur mustard (SM) is a chemical warfare agent that can damage DNA via alkylation and oxidative stress. Because of its genotoxicity, SM is cancerogenic and the progenitor of many chemotherapeutics. Previously, we developed an SM-resistant cell line via chronic exposure of the popular keratinocyte cell line HaCaT to increasing doses of SM over a period of 40 months. In this study, we compared the genomic landscape of the SM-resistant cell line HaCaT/SM to its sensitive parental line HaCaT in order to gain insights into genetic changes associated with continuous alkylation and oxidative stress. We established chromosome numbers by cytogenetics, analyzed DNA copy number changes by means of array Comparative Genomic Hybridization (array CGH), employed the genome-wide chromosome conformation capture technique Hi-C to detect chromosomal translocations, and derived mutational signatures by whole-genome sequencing. We observed that chronic SM exposure eliminated the initially prevailing hypotetraploid cell population in favor of a hyperdiploid one, which contrasts with previous observations that link polyploidization to increased tolerance and adaptability toward genotoxic stress. Furthermore, we observed an accumulation of chromosomal translocations, frequently flanked by DNA copy number changes, which indicates a high rate of DNA double-strand breaks and their misrepair. HaCaT/SM-specific single-nucleotide variants showed enrichment of C > A and T > A transversions and a lower rate of deaminated cytosines in the CpG dinucleotide context. Given the frequent use of HaCaT in toxicology, this study provides a valuable data source with respect to the original genotype of HaCaT and the mutational signatures associated with chronic alkylation and oxidative stress.

A Novel Locus and Candidate Gene for Familial Developmental Dyslexia on Chromosome 4q.

Objective: Developmental dyslexia is a highly heritable specific reading and writing disability. To identify a possible new locus and candidate gene for this disability, we investigated a four-generation pedigree where transmission of dyslexia is consistent with an autosomal dominant inheritance pattern. Methods: We performed genome wide array-based SNP genotyping and parametric linkage analysis and sequencing analysis of protein-coding exons, exon-intron boundaries and conserved extragenic regions within the haplotype cosegregating with dyslexia in DNA from one affected and one unaffected family member. Cosegregation was confirmed by sequencing all available family members. Additionally, we analyzed 96 dyslexic individuals who had previously shown positive LOD scores on chromosome 4q28 as well as an even larger sample (n = 2591). Results: We found a single prominent linkage interval on chromosome 4q, where sequence analysis revealed a nucleotide variant in the 3' UTR of brain expressed SPRY1 in the dyslexic family member that cosegregated with dyslexia. This sequence alteration might affect the binding efficiency of the IGF2BP1 RNA-binding protein and thus influence the expression level of the SPRY1 gene product. An analysis of 96 individuals from a cohort of dyslexic individuals revealed a second heterozygous variant in this gene, which was absent in the unaffected sister of the proband. An investigation of the region in a much larger sample further found a nominal p-value of 0.0016 for verbal short-term memory (digit span) in 2,591 individuals for a neighboring SNV. After correcting for the local number of analyzed SNVs, and after taking into account linkage disequilibrium, we found this corresponds to a p-value of 0.0678 for this phenotype. Conclusions: We describe a new locus for familial dyslexia and discuss the possibility that SPRY1 might play a role in the etiology of a monogenic form of dyslexia.

CT Irradiation-induced Changes of Gene Expression within Peripheral Blood Cells.

Computed tomography (CT) is a crucial element of medical imaging diagnostics. The widespread application of this technology has made CT one of the major contributors to medical radiation burden, despite the fact that doses per individual CT scan steadily decrease due to the advancement of technology. Epidemiological risk assessment of CT exposure is hampered by the fact that moderate adverse effects triggered by low doses of CT exposure are likely masked by statistical fluctuations. In light of these limitations, there is need of further insights into the biological processes induced by CT scans to complement the existing knowledge base of risk assessment. This prompted us to investigate the early transcriptomic response of ex vivo irradiated peripheral blood of three healthy individuals. Samples were irradiated employing a modern dual-source-CT-scanner with a tube voltage of 150 kV, resulting in an estimated effective dose of 9.6 mSv. RNA was isolated 1 h and 6 h after exposure, respectively, and subsequently analyzed by RNA deep sequencing. Differential gene expression analysis revealed shared upregulation of AEN, FDXR, and DDB2 6 h after exposure in all three probands. All three genes have previously been discussed as radiation responsive genes and have already been implicated in DNA damage response and cell cycle control after DNA damage. In summary, we substantiated the usefulness of AEN, FDXR, and DDB2 as RNA markers of low dose irradiation. Moreover, the upregulation of genes associated with DNA damage reminds one of the genotoxic nature of CT diagnostics even with the low doses currently applied.

X-ray irradiation induces subtle changes in the genome-wide distribution of DNA hydroxymethylation with opposing trends in genic and intergenic regions.

DNA hydroxymethylation has gained attention as an intermediate in the process of DNA demethylation. More recently, 5-hydroxymethylcytosine has been recognized as an independent epigenetic mark that can persist over time and that exerts influence on gene regulation and other biological processes. Deregulation of this DNA modification has been linked to tumorigenesis and a variety of other diseases. The impact of irradiation on DNA hydroxymethylation is poorly understood. In this study we exposed lung fibroblasts (IMR90) to 0.5 Gy and 2 Gy of X-rays, respectively. We characterized radiation induced changes of DNA hydroxymethylation 1 h, 6 h, 24 h and 120 h after exposure employing immunoprecipitation and subsequent deep sequencing of the genomic fraction enriched for hydroxymethylated DNA. Transcriptomic response to irradiation was analyzed for time points 6 h and 24 h post exposure by means of RNA sequencing. Irradiated and sham-irradiated samples shared the same overall distribution of 5-hydroxymethylcytosines with respect to genomic features such as promoters and exons. The frequency of 5-hydroxymethylcytosine peaks differentially detected in irradiated samples increased in genic regions over time, while the opposing trend was observed for intergenic regions. Onset and extent of this effect was dose dependent. Moreover, we demonstrated a biased distribution of 5-hmC alterations at CpG islands and sites occupied by the DNA binding protein CTCF. In summary, our study provides new insights into the epigenetic response to irradiation. Our data highlight genomic features more prone to irradiation induced changes of DNA hydroxymethylation, which might impact early and late onset effects of irradiation.

Gene expression changes in human iPSC-derived cardiomyocytes after X-ray irradiation.

Purpose: Radiation-induced heart disease caused by cardiac exposure to ionizing radiation comprises a variety of cardiovascular effects. Research in this field has been hampered by limited availability of clinical samples and appropriate test models. In this study, we wanted to elucidate the molecular mechanisms underlying electrophysiological changes, which we have observed in a previous study. Materials and methods: We employed RNA deep-sequencing of human-induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) 48 h after 5 Gy X-ray irradiation. By comparison to public data from hiPSC-CMs and human myocardium, we verified the expression of cardiac-specific genes in hiPSC-CMs. Results were validated by qRT-PCR. Results: Differentially gene expression analysis identified 39 and 481 significantly up- and down-regulated genes after irradiation, respectively. Besides, a large fraction of genes associated with cell cycle processes, we identified genes implicated in cardiac calcium homeostasis (PDE3B), oxidative stress response (FDXR and SPATA18) and the etiology of cardiomyopathy (SGCD, BBC3 and GDF15). Conclusions: Notably, observed gene expression characteristics specific to hiPSC-CMs might be relevant regarding further investigations of the response to external stressors like radiation. The genes and biological processes highlighted in our study present promising starting points for functional follow-up studies for which hiPSC-CMs could pose an appropriate cell model when cell type specific peculiarities are taken into account.

Genome-Wide Analysis of Interchromosomal Interaction Probabilities Reveals Chained Translocations and Overrepresentation of Translocation Breakpoints in Genes in a Cutaneous T-Cell Lymphoma Cell Line.

In classical models of tumorigenesis, the accumulation of tumor promoting chromosomal aberrations is described as a gradual process. Next-generation sequencing-based methods have recently revealed complex patterns of chromosomal aberrations, which are beyond explanation by these classical models of karyotypic evolution of tumor genomes. Thus, the term chromothripsis has been introduced to describe a phenomenon, where temporarily and spatially confined genomic instability results in dramatic chromosomal rearrangements limited to segments of one or a few chromosomes. Simultaneously arising and misrepaired DNA double-strand breaks are also the cause of another phenomenon called chromoplexy, which is characterized by the presence of chained translocations and interlinking deletion bridges involving several chromosomes. In this study, we demonstrate the genome-wide identification of chromosomal translocations based on the analysis of translocation-associated changes in spatial proximities of chromosome territories on the example of the cutaneous T-cell lymphoma cell line Se-Ax. We have used alterations of intra- and interchromosomal interaction probabilities as detected by genome-wide chromosome conformation capture (Hi-C) to infer the presence of translocations and to fine-map their breakpoints. The outcome of this analysis was subsequently compared to datasets on DNA copy number alterations and gene expression. The presence of chained translocations within the Se-Ax genome, partly connected by intervening deletion bridges, indicates a role of chromoplexy in the etiology of this cutaneous T-cell lymphoma. Notably, translocation breakpoints were significantly overrepresented in genes, which highlight gene-associated biological processes like transcription or other gene characteristics as a possible cause of the observed complex rearrangements. Given the relevance of chromosomal aberrations for basic and translational research, genome-wide high-resolution analysis of structural chromosomal aberrations will gain increasing importance.

Impact of Ionizing Radiation on Electrophysiological Behavior of Human-induced Ipsc-derived Cardiomyocytes on Multielectrode Arrays.

Cardiac arrhythmia presumably induced through cardiac fibrosis is a recurrent long-term consequence of exposure to ionizing radiation. However, there is also evidence that cardiac arrhythmia can occur in patients shortly after irradiation. In this study, the authors employed multielectrode arrays to investigate the short-term effects of x-ray radiation on the electrophysiological behavior of cardiomyocytes derived from human-induced pluripotent stem cells. These cardiomyocytes with spontaneous pacemaker activity were cultured on single-well multielectrode arrays. After exposure to 0, 0.5, 1, 2, 5, 10 Gy x-ray radiation, electrical activity was measured at time points ranging from 10 min to 96 h. RNA sequencing was employed to verify the expression of genes specifically involved in cardiomyocyte differentiation and function. A decrease in beating rate was observed after irradiation with 5 and 10 Gy starting 48 h after exposure. Cells exposed to higher doses of radiation were more prone to show changes in electrophysiological spatial distribution. No radiation-induced effects with respect to the corrected QT interval were detectable. Gene expression analysis showed up regulation of typical cardiac features like ACTC1 or HCN4. In this study, early dose-dependent changes in electrophysiological behavior were determined after x-ray irradiation. Results point towards a dose-dependent effect on pacemaker function of cardiomyocytes and indicate a possible connection between irradiation and short-term changes in electrophysiological cardiac function. Cardiomyocytes derived from human-induced pluripotent stem cells on multielectrode arrays represent a promising in vitro cardiac-modeling system for preclinical studies.

Gene Expression Analysis in Human Peripheral Blood Cells after 900 MHz RF-EMF Short-Term Exposure.

Radiofrequency electromagnetic fields (RF-EMF) are a basic requirement of modern wireless communication technology. Statutory thresholds of RF-EMF are established to limit relevant additional heat supply in human tissue. Nevertheless, to date, questions concerning nonthermal biological effects have yet to be fully addressed. New versions of microarrays (8 × 60K v2) provide a higher resolution of whole genome gene expression to display adaptive processes in cells after irradiation. In this ex vivo/ in vitro study, we irradiated peripheral blood cells from five donors with a continuous wave of 900 MHz RF-EMF for 0, 30, 60 and 90 min. Gene expression changes ( P ≤ 0.05 and ≥twofold differences above or below the room temperature control exposed samples) were evaluated with microarray analysis. The results were compared with data from room temperature + 2°C samples. Verification of microarray results was performed using bioinformatic analyses and qRT-PCR. We registered a lack of an EMF-specific gene expression response after applying the false discovery rate adjustment (FDR), using a high-stringency approach. Low-stringency analysis revealed 483 statistically significant deregulated transcripts in all RF-EMF groups relative to the room temperature exposed samples without an association with their corresponding room temperature + 2°C controls. Nevertheless, these transcripts must be regarded as statistical artefacts due to the absence of a targeted biological response, including enrichment and network analyses administered to microarray expressed gene subset profiles. Correspondingly, 14 most promising candidate transcripts examined by qRT-PCR displayed an absence of correlation with respect to the microarray results. In conclusion, these findings indicate that 900 MHz EMF exposure establishing an average specific absorption rate of 9.3 W/kg to whole blood cells is insufficient to induce nonthermal effects in gene expression during short-time exposure up to 90 min.

Generation of a human induced pluripotent stem cell (iPSC) line from a 51-year-old female with attention-deficit/hyperactivity disorder (ADHD) carrying a duplication of SLC2A3.

Fibroblasts were isolated from a skin biopsy of a clinically diagnosed 51-year-old female attention-deficit/hyperactivity disorder (ADHD) patient carrying a duplication of SLC2A3, a gene encoding neuronal glucose transporter-3 (GLUT3). Patient fibroblasts were infected with Sendai virus, a single-stranded RNA virus, to generate transgene-free human induced pluripotent stem cells (iPSCs). SLC2A3-D2-iPSCs showed expression of pluripotency-associated markers, were able to differentiate into cells of the three germ layers in vitro and had a normal female karyotype. This in vitro cellular model can be used to study the role of risk genes in the pathogenesis of ADHD, in a patient-specific manner.

Exploring the Link between Radiation Exposure and Multifocal Basal Cell Carcinomas in a Former Chernobyl Clean-up Worker by Combining Different Molecular Biological Techniques.

Thirty years after the Chernobyl nuclear power plant accident we report on a patient who was a clean-up worker, who subsequently developed multiple cutaneous basal cell carcinomas (BCCs). We used several methods to assess the biological long-term effects related to low-dose external and internal radiation exposure. Specifically, because BCC risk may be increased with ionizing radiation exposure, we endeavored to determine whether the multifocal BCCs were related to the patient's past clean-up work. We assessed cytogenetic changes using peripheral blood, and internal incorporation was measured with a whole-body counter. Gene expression alterations were determined and array-based comparative genomic hybridization was performed for copy number aberration analysis of available BCC samples. In 1,053 metaphase cells, the dicentric yield of 0.005 dicentrics, with acentrics/cell, was significantly increased compared to the established calibration curve (P < 0.001). A 2.5-fold increase in total translocations was observed compared to the expected translocation rate. No internal contamination was detected with the whole-body counter. At the RNA level, two of seven genes (HNRNPA1, AGAP4/6/8) indicated internal plutonium exposure associated with the lowest dose category found in Mayak workers (>0-0.055 Gy). Relevant DNA copy number changes were only detected within the most aggressive BCC focus. Our results suggest that the examined worker had low and more recent radiation exposure with presumably internalized radionuclides that were below the detection level of a whole-body counter. The multifocal BCC could not be related to past occupational radiation exposure. The findings from our study suggest that integrating different methodologies potentially provides an improved overall assessment of individual health risks associated with or excluding occupational radiation exposure.

MEF2C-dysregulated pediatric T-cell acute lymphoblastic leukemia is associated with CDKN1B deletions and a poor response to glucocorticoid therapy.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological disease in which multiple genetic abnormalities cooperate in the malignant transformation of T-lymphoid progenitors. Although in pediatric T-ALL, CDKN1B deletions occur in about 12% of the cases and represent one of the most frequent copy number alterations, neither their association with other genetic alterations nor the clinical characteristics of these patients have been determined yet. In this study, we show that loss of CDKN1B increased the prevalence of cell cycle regulator defects in immature T-ALL, usually only ascribed to CDKN2A/B deletions, and that CDKN1B deletions frequently coincide with expression of MEF2C, considered as one of the driving oncogenes in immature early T-cell precursor (ETP) ALL. However, MEF2C-dysregulation was only partially associated with the immunophenotypic characteristics used to define ETP-ALL. Furthermore, MEF2C expression levels were significantly associated with or may even be predictive of the response to glucocorticoid treatment.

Pre-Exposure Gene Expression in Baboons with and without Pancytopenia after Radiation Exposure.

Radiosensitivity differs in humans and likely among primates. The reasons are not well known. We examined pre-exposure gene expression in baboons (n = 17) who developed haematologic acute radiation syndrome (HARS) without pancytopenia or a more aggravated HARS with pancytopenia after irradiation. We evaluated gene expression in a two stage study design where stage I comprised a whole genome screen for messenger RNAs (mRNA) (microarray) and detection of 667 microRNAs (miRNA) (real-time quantitative polymerase chain reaction (qRT-PCR) platform). Twenty candidate mRNAs and nine miRNAs were selected for validation in stage II (qRT-PCR). None of the mRNA species could be confirmed during the validation step, but six of the nine selected candidate miRNA remained significantly different during validation. In particular, miR-425-5p (receiver operating characteristic = 0.98; p = 0.0003) showed nearly complete discrimination between HARS groups with and without pancytopenia. Target gene searches of miR-425-5p identified new potential mRNAs and associated biological processes linked with radiosensitivity. We found that one miRNA species examined in pre-exposure blood samples was associated with HARS characterized by pancytopenia and identified new target mRNAs that might reflect differences in radiosensitivity of irradiated normal tissue.

SLC2A3 single-nucleotide polymorphism and duplication influence cognitive processing and population-specific risk for attention-deficit/hyperactivity disorder.

BACKGROUND: Attention-deficit/hyperactivity disorder (ADHD) is a common, highly heritable neurodevelopmental disorder with profound cognitive, behavioral, and psychosocial impairments with persistence across the life cycle. Our initial genome-wide screening approach for copy number variants (CNVs) in ADHD implicated a duplication of SLC2A3, encoding glucose transporter-3 (GLUT3). GLUT3 plays a critical role in cerebral glucose metabolism, providing energy for the activity of neurons, which, in turn, moderates the excitatory-inhibitory balance impacting both brain development and activity-dependent neural plasticity. We therefore aimed to provide additional genetic and functional evidence for GLUT3 dysfunction in ADHD. METHODS: Case-control association analyses of SLC2A3 single-nucleotide polymorphisms (SNPs) and CNVs were conducted in several European cohorts of patients with childhood and adult ADHD (SNP, n = 1,886 vs. 1,988; CNV, n = 1,692 vs. 1,721). These studies were complemented by SLC2A3 expression analyses in peripheral cells, functional EEG recordings during neurocognitive tasks, and ratings of food energy content. RESULTS: Meta-analysis of all cohorts detected an association of SNP rs12842 with ADHD. While CNV analysis detected a population-specific enrichment of SLC2A3 duplications only in German ADHD patients, the CNV + rs12842 haplotype influenced ADHD risk in both the German and Spanish cohorts. Duplication carriers displayed elevated SLC2A3 mRNA expression in peripheral blood cells and altered event-related potentials reflecting deficits in working memory and cognitive response control, both endophenotypic traits of ADHD, and an underestimation of energy units of high-caloric food. CONCLUSIONS: Taken together, our results indicate that both common and rare SLC2A3 variation impacting regulation of neuronal glucose utilization and energy homeostasis may result in neurocognitive deficits known to contribute to ADHD risk.

GenomeCAT: a versatile tool for the analysis and integrative visualization of DNA copy number variants.

BACKGROUND: The analysis of DNA copy number variants (CNV) has increasing impact in the field of genetic diagnostics and research. However, the interpretation of CNV data derived from high resolution array CGH or NGS platforms is complicated by the considerable variability of the human genome. Therefore, tools for multidimensional data analysis and comparison of patient cohorts are needed to assist in the discrimination of clinically relevant CNVs from others. RESULTS: We developed GenomeCAT, a standalone Java application for the analysis and integrative visualization of CNVs. GenomeCAT is composed of three modules dedicated to the inspection of single cases, comparative analysis of multidimensional data and group comparisons aiming at the identification of recurrent aberrations in patients sharing the same phenotype, respectively. Its flexible import options ease the comparative analysis of own results derived from microarray or NGS platforms with data from literature or public depositories. Multidimensional data obtained from different experiment types can be merged into a common data matrix to enable common visualization and analysis. All results are stored in the integrated MySQL database, but can also be exported as tab delimited files for further statistical calculations in external programs. CONCLUSIONS: GenomeCAT offers a broad spectrum of visualization and analysis tools that assist in the evaluation of CNVs in the context of other experiment data and annotations. The use of GenomeCAT does not require any specialized computer skills. The various R packages implemented for data analysis are fully integrated into GenomeCATs graphical user interface and the installation process is supported by a wizard. The flexibility in terms of data import and export in combination with the ability to create a common data matrix makes the program also well suited as an interface between genomic data from heterogeneous sources and external software tools. Due to the modular architecture the functionality of GenomeCAT can be easily extended by further R packages or customized plug-ins to meet future requirements.

MicroRNA Expression for Early Prediction of Late Occurring Hematologic Acute Radiation Syndrome in Baboons.

For effective medical management of radiation-exposed persons after a radiological/nuclear event, blood-based screening measures in the first few days that could predict hematologic acute radiation syndrome (HARS) are needed. For HARS severity prediction, we used microRNA (miRNA) expression changes measured on days one and two after irradiation in a baboon model. Eighteen baboons underwent different patterns of partial or total body irradiation, corresponding to an equivalent dose of 2.5 or 5 Gy. According to changes in blood cell counts (BCC) the surviving baboons (n = 17) exhibited mild (H1-2, n = 4) or more severe (H2-3, n = 13) HARS. In a two Stage study design we screened 667 miRNAs using a quantitative real-time polymerase chain reaction (qRT-PCR) platform. In Stage II we validated candidates where miRNAs had to show a similar regulation (up- or down-regulated) and a significant 2-fold miRNA expression difference over H0. Seventy-two candidate miRNAs (42 for H1-2 and 30 for H2-3) were forwarded for validation. Forty-two of the H1-2 miRNA candidates from the screening phase entered the validation step and 20 of them showed a statistically significant 2-4 fold up-regulation relative to the unexposed reference (H0). Fifteen of the 30 H2-3 miRNAs were validated in Stage II. All miRNAs appeared 2-3 fold down-regulated over H0 and allowed an almost complete separation of HARS categories; the strongest candidate, miR-342-3p, showed a sustained and 10-fold down-regulation on both days 1 and 2. In summary, our data support the medical decision making of the HARS even within the first two days after exposure where diagnostic tools for early medical decision are required but so far missing. The miRNA species identified and in particular miR-342-3p add to the previously identified mRNAs and complete the portfolio of identified mRNA and miRNA transcripts for HARS prediction and medical management.

Inactivation of RUNX3/p46 Promotes Cutaneous T-Cell Lymphoma.

The key role of RUNX3 in physiological T-cell differentiation has been extensively documented. However, information on its relevance for the development of human T-cell lymphomas or leukemias is scarce. Here, we show that alterations of RUNX3 by either heterozygous deletion or methylation of its distal promoter can be observed in the tumor cells of 15 of 21 (71%) patients suffering from Sézary syndrome, an aggressive variant of cutaneous T-cell lymphoma. As a consequence, mRNA levels of RUNX3/p46, the isoform controlled by the distal promoter, are significantly lower in Sézary syndrome tumor cells. Re-expression of RUNX3/p46 reduces cell viability and promotes apoptosis in a RUNX3/p46low cell line of cutaneous T-cell lymphoma. Based on this, we present evidence that RUNX3 can act as a tumor suppressor in a human T-cell malignancy and suggest that this effect is predominantly mediated through transcripts from its distal promoter, in particular RUNX3/p46.