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Objective: To prospectively investigate the influence of contrast material concentration on enhancement in cardiac CT by using a biphasic single-injection protocol.Methods: Sixty-four-row multidetector cardiac CT angiography was performed in 159 patients randomised to a moderate or high contrast medium concentration. Contrast material injection included a first phase for enhancement of the coronary arteries and a second phase, at half the iodine flux, targeted at enhancement of the right ventricle. Contrast medium injection was followed by a saline flush. For both concentrations, injection duration (and thus total iodine dose) was adapted to the duration of the CT data acquisition and iodine flux was adjusted to patient weight. Attenuation was measured at various levels in the heart and vessels and the two concentrations compared, overall and per weight group.Results: Enhancement of the aorta and left ventricle was significantly greater with the moderate than with the high concentration contrast medium. This remained true for the two higher weight groups. No difference was found in the lowest weight group or in the right ventricle and pulmonary outflow tract.Conclusion: With a biphasic injection protocol, enhancement of the aorta and left ventricle was weaker with the higher concentration of contrast material. [ABSTRACT FROM AUTHOR]

Background: Coronary calcification can lead to over-estimation of the degree of coronary stenosis. Purpose: To evaluate whether thinner reconstruction thickness improves the diagnostic performance of 64-slice CT coronary angiography (CTCA) in angina patients with a positive calcium score. Material and Methods: We selected 20 scans from a clinical study comparing CTCA to conventional coronary angiography (CCA) in stable and unstable angina patients based on a low number of motion artifacts and a positive calcium score. All images were acquired at 64 x 0.625 mm and each CTCA scan was reconstructed at slice thickness/increment 0.67 mm/0.33 mm, 0.9 mm/0.45 mm, and 1.4 mm/0.7 mm. Two reviewers blinded for CCA results independently evaluated the scans for the presence of significant coronary artery disease (CAD) in three randomly composed series, with > or =2 weeks in between series. The diagnostic performance of CTCA was compared for the different slice thicknesses using a pooled analysis of both reviewers. Significant CAD was defined as >50% diameter narrowing on quantitative CCA. Image noise (standard deviation of CT numbers) was measured in all scans. Inter-observer variability was assessed with kappa. Results: Significant CAD was present in 8% of 304 available segments. Median total Agatston calcium score was 181.8 (interquartile range 34.9-815.6). Sensitivity at 0.67 mm, 0.9 mm, and 1.4 mm slice thickness was 70% (95% confidence interval 57-83%), 74% (62-86%), and 70% (57-83%), respectively. Specificity was 85% (82-88%), 84% (81-87%), and 84% (81-87%), respectively. The positive predictive value was 30 (21-38%), 29 (21-37%), and 28 (20-36%), respectively. The negative predictive value was 97% (95-98%), 97% (96-99%), and 97% (96-99%), respectively. Kappa for inter-observer agreement was 0.56, 0.58, and 0.59. Noise decreased from 32.9 HU at 0.67 mm, to 23.2 HU at 1.4 mm (P<0.001). Conclusion: Diagnostic performance of CTCA in angina patients with a positive calcium score was not markedly affected by modest variations in reconstruction slice thickness. [ABSTRACT FROM AUTHOR]

Purpose: To utilize 40-slice electrocardiographically (ECG)-gated cine computed tomographic angiography (CTA) to characterize normal aortic motion during the cardiac cycle at relevant anatomical landmarks in preoperative abdominal aortic aneurysm (AAA) patients. Methods: In 10 consecutive preoperative AAA patients (10 men; mean age 78.8 years, range 69-86), an ECG-gated CTA dataset was acquired on a 40-slice CT scanner using a standard radiation dose. CTA quality was graded and scan time was measured. Pulsatility measurements at multiple relevant anatomical levels were performed in the axial plane. Changes in aortic circumference were determined for both the aortic wall and the luminal diameter. Results: All 10 CT scans were of good quality. All patients could be scanned in 14 to 33 seconds (mean 21). At each anatomical level measured, there was a 2.2- to 3.4-mm increase in the aortic wall circumference per cardiac cycle. A similar increase was observed in luminal circumference, with a 2.4- to 3.6-mm increase per cycle. Conclusion: This study introduces the concept of dynamic cine CTA imaging of aortic motion, providing insight into the pathophysiology of abdominal aortic and iliac pulsations. Patients with AAAs selected for EVAR demonstrate changes in aortic circumference with each cardiac cycle that may have consequences for endograft sizing and future design. The potential for graft migration, intermittent type I endoleak, and poor patient outcome following EVAR can be anticipated. Complex aortic dynamics deserve increased scrutiny in an effort to prevent potential complications. [ABSTRACT FROM AUTHOR]

The biological mechanisms and potential clinical impact of heterotopic ossification (HO) in colorectal neoplasms are not fully understood. This study investigates the clinicopathological characteristics of colorectal neoplasms associated with HO and examines the potential role of the bone morphogenetic protein (BMP) pathway in development of HO. An artificial intelligence (AI) based classification of colorectal cancers (CRC) exhibiting HO and their association with consensus molecular subtypes (CMS) is performed. The study included 77 cases via the Dutch nationwide Pathology databank. Immunohistochemistry for BMP2, SMAD4, and Osterix was performed. An AI algorithm assessed the tumour-stroma ratio to approximate the CMS. A literature search yielded 96 case reports, which were analysed and compared with our cases for clinicopathological parameters. HO was more frequently observed in our cohort in traditional serrated adenomas (25%), tubulovillous adenomas (25%) and juvenile polyps (25%), while in the literature it was most often seen in juvenile polyps (38.2%) and inflammatory polyps (29.4%). In both cohorts, carcinomas were mostly conventional (>60%) followed by mucinous and serrated adenocarcinomas. Higher expression of BMP2, SMAD4, and Osterix was observed in tumour and/or stromal cells directly surrounding bone, indicating activation of the BMP pathway. The tumour-stroma analysis appointed >50% of the cases to the mesenchymal subtype (CMS4) (59%). HO has a predilection for serrated and juvenile/inflammatory polyps, mucinous and serrated adenocarcinomas. BMP signalling is activated and seems to play a role in formation of HO in colorectal neoplasms. In line with TGFβ/BMP pathway activation associated with CMS4 CRC, HO seems associated with CMS4., Competing Interests: Declaration of competing interest No potential conflicts of interest relevant to this article were reported., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Oesophageal adenocarcinomas may show different histopathological patterns, including excessive acellular mucin pools, signet-ring cells (SRCs), and poorly cohesive cells (PCCs). These components have been suggested to correlate with poor outcomes after neoadjuvant chemoradiotherapy (nCRT), which might influence patient management. However, these factors have not been studied independently of each other with adjustment for tumour differentiation grade (i.e. the presence of well-formed glands), which is a possible confounder. We studied the pre- and post-treatment presence of extracellular mucin, SRCs, and/or PCCs in relation to pathological response and prognosis after nCRT in patients with oesophageal or oesophagogastric junction adenocarcinoma. A total of 325 patients were retrospectively identified from institutional databases of two university hospitals. All patients were scheduled for ChemoRadiotherapy for Oesophageal cancer followed by Surgery Study (CROSS) nCRT and oesophagectomy between 2001 and 2019. Percentages of well-formed glands, extracellular mucin, SRCs, and PCCs were scored in pre-treatment biopsies and post-treatment resection specimens. The association between histopathological factors (≥1 and >10%) and tumour regression grade 3-4 (i.e. >10% residual tumour), overall survival, and disease-free survival (DFS) was evaluated, adjusted for tumour differentiation grade amongst other clinicopathological variables. In pre-treatment biopsies, ≥1% extracellular mucin was present in 66 of 325 patients (20%); ≥1% SRCs in 43 of 325 (13%), and ≥1% PCCs in 126 of 325 (39%). We show that pre-treatment histopathological factors were unrelated to tumour regression grade. Pre-treatment presence of >10% PCCs was associated with lower DFS (hazard ratio [HR] 1.73, 95% CI 1.19-2.53). Patients with post-treatment presence of ≥1% SRCs had higher risk of death (HR 1.81, 95% CI 1.10-2.99). In conclusion, pre-treatment presence of extracellular mucin, SRCs, and/or PCCs is unrelated to pathological response. The presence of these factors should not be an argument to refrain from CROSS. At least 10% PCCs pre-treatment and any SRCs post-treatment, irrespective of the tumour differentiation grade, seem indicative of inferior prognosis, but require further validation in larger cohorts., (© 2023 The Authors. The Journal of Pathology: Clinical Research published by The Pathological Society of Great Britain and Ireland and John Wiley & Sons Ltd.)

Analysis of predicted fungal proteomes revealed a large family of sequences that showed similarity to the Saccharomyces cerevisiae Class-I dihydroorotate dehydrogenase Ura1, which supports synthesis of pyrimidines under aerobic and anaerobic conditions. However, expression of codon-optimised representatives of this gene family, from the ascomycete Alternaria alternata and the basidiomycete Schizophyllum commune, only supported growth of an S. cerevisiae ura1Δ mutant when synthetic media were supplemented with dihydrouracil. A hypothesis that these genes encode NAD(P) + -dependent dihydrouracil dehydrogenases (EC 1.3.1.1 or 1.3.1.2) was rejected based on absence of complementation in anaerobic cultures. Uracil- and thymine-dependent oxygen consumption and hydrogen-peroxide production by cell extracts of S. cerevisiae strains expressing the A. alternata and S. commune genes showed that, instead, they encode active dihydrouracil oxidases (DHO, EC1.3.3.7). DHO catalyses the reaction dihydrouracil + O 2  → uracil + H 2 O 2 and was only reported in the yeast Rhodotorula glutinis (Owaki in J Ferment Technol 64:205-210, 1986). No structural gene for DHO was previously identified. DHO-expressing strains were highly sensitive to 5-fluorodihydrouracil (5F-dhu) and plasmids bearing expression cassettes for DHO were readily lost during growth on 5F-dhu-containing media. These results show the potential applicability of fungal DHO genes as counter-selectable marker genes for genetic modification of S. cerevisiae and other organisms that lack a native DHO. Further research should explore the physiological significance of this enigmatic and apparently widespread fungal enzyme., (© 2022. The Author(s).)

Introduction: Patients with coronavirus disease 2019 (COVID-19) can present with chest pain. However, the characteristics of this chest pain are unknown. We performed a single-centre observational study to review and summarise chest pain characteristics in COVID-19 patients at first presentation to the emergency department (ED)., Methods: We collected data on characteristics of 'chest pain' reported by COVID-19 patients who attended the ED of Bernhoven Hospital, the Netherlands from 4 through 30 March 2020., Results: We included 497 COVID-19 patients, of whom 83 (17%) reported chest pain upon presentation to the ED. Chest pain characteristics were: present since disease onset (88%), retrosternal location (43%), experienced as compressing/pressure pain (61%), no radiation (61%) and linked to heavy coughing (39%). Patients who reported chest pain were younger than those without chest pain (61 vs 73 years; p < 0.001). Patients with syncope were older (75 vs 72 years; p = 0.017), had a shorter duration of symptoms (5 vs 7 days; p < 0.001) and reported fewer respiratory complaints (68% vs 90%; p < 0.001) than those without syncope. Patients with new-onset atrial arrhythmias presented with a shorter duration of symptoms (5 vs 7 days; p = 0.013), experienced fewer respiratory complaints (72% vs 89%; p = 0.012) and more frequently had a history of cardiovascular disease (79% vs 50%; p = 0.003) than patients who presented without arrythmias., Conclusion: Chest pain and other cardiac symptoms were frequently observed in COVID-19 patients. Treating physicians should be aware that chest pain, arrhythmias and syncope can be presenting symptoms of COVID-19., (© 2022. The Author(s).)

Background: In most fungi, quinone-dependent Class-II dihydroorotate dehydrogenases (DHODs) are essential for pyrimidine biosynthesis. Coupling of these Class-II DHODHs to mitochondrial respiration makes their in vivo activity dependent on oxygen availability. Saccharomyces cerevisiae and closely related yeast species harbor a cytosolic Class-I DHOD (Ura1) that uses fumarate as electron acceptor and thereby enables anaerobic pyrimidine synthesis. Here, we investigate DHODs from three fungi (the Neocallimastigomycete Anaeromyces robustus and the yeasts Schizosaccharomyces japonicus and Dekkera bruxellensis) that can grow anaerobically but, based on genome analysis, only harbor a Class-II DHOD., Results: Heterologous expression of putative Class-II DHOD-encoding genes from fungi capable of anaerobic, pyrimidine-prototrophic growth (Arura9, SjURA9, DbURA9) in an S. cerevisiae ura1Δ strain supported aerobic as well as anaerobic pyrimidine prototrophy. A strain expressing DbURA9 showed delayed anaerobic growth without pyrimidine supplementation. Adapted faster growing DbURA9-expressing strains showed mutations in FUM1, which encodes fumarase. GFP-tagged SjUra9 and DbUra9 were localized to S. cerevisiae mitochondria, while ArUra9, whose sequence lacked a mitochondrial targeting sequence, was localized to the yeast cytosol. Experiments with cell extracts showed that ArUra9 used free FAD and FMN as electron acceptors. Expression of SjURA9 in S. cerevisiae reproducibly led to loss of respiratory competence and mitochondrial DNA. A cysteine residue (C265 in SjUra9) in the active sites of all three anaerobically active Ura9 orthologs was shown to be essential for anaerobic activity of SjUra9 but not of ArUra9., Conclusions: Activity of fungal Class-II DHODs was long thought to be dependent on an active respiratory chain, which in most fungi requires the presence of oxygen. By heterologous expression experiments in S. cerevisiae, this study shows that phylogenetically distant fungi independently evolved Class-II dihydroorotate dehydrogenases that enable anaerobic pyrimidine biosynthesis. Further structure-function studies are required to understand the mechanistic basis for the anaerobic activity of Class-II DHODs and an observed loss of respiratory competence in S. cerevisiae strains expressing an anaerobically active DHOD from Sch. japonicus., (© 2021. The Author(s).)

Neocallimastigomycetes are unique examples of strictly anaerobic eukaryotes. This study investigates how these anaerobic fungi bypass reactions involved in synthesis of pyridine nucleotide cofactors and coenzyme A that, in canonical fungal pathways, require molecular oxygen. Analysis of Neocallimastigomycetes proteomes identified a candidate l-aspartate-decarboxylase (AdcA) and l-aspartate oxidase (NadB) and quinolinate synthase (NadA), constituting putative oxygen-independent bypasses for coenzyme A synthesis and pyridine nucleotide cofactor synthesis. The corresponding gene sequences indicated acquisition by ancient horizontal gene transfer (HGT) events involving bacterial donors. To test whether these enzymes suffice to bypass corresponding oxygen-requiring reactions, they were introduced into fms1 Δ and bna2 Δ Saccharomyces cerevisiae strains. Expression of nadA and nadB from Piromyces finnis and adcA from Neocallimastix californiae conferred cofactor prototrophy under aerobic and anaerobic conditions. This study simulates how HGT can drive eukaryotic adaptation to anaerobiosis and provides a basis for elimination of auxotrophic requirements in anaerobic industrial applications of yeasts and fungi. IMPORTANCE NAD (NAD + ) and coenzyme A (CoA) are central metabolic cofactors whose canonical biosynthesis pathways in fungi require oxygen. Anaerobic gut fungi of the Neocallimastigomycota phylum are unique eukaryotic organisms that adapted to anoxic environments. Analysis of Neocallimastigomycota genomes revealed that these fungi might have developed oxygen-independent biosynthetic pathways for NAD + and CoA biosynthesis, likely acquired through horizontal gene transfer (HGT) from prokaryotic donors. We confirmed functionality of these putative pathways under anaerobic conditions by heterologous expression in the yeast Saccharomyces cerevisiae. This approach, combined with sequence comparison, offers experimental insight on whether HGT events were required and/or sufficient for acquiring new traits. Moreover, our results demonstrate an engineering strategy for enabling S. cerevisiae to grow anaerobically in the absence of the precursor molecules pantothenate and nicotinate, thereby contributing to alleviate oxygen requirements and to move closer to prototrophic anaerobic growth of this industrially relevant yeast.

The Gleason score is the most important prognostic marker for prostate cancer patients, but it suffers from significant observer variability. Artificial intelligence (AI) systems based on deep learning can achieve pathologist-level performance at Gleason grading. However, the performance of such systems can degrade in the presence of artifacts, foreign tissue, or other anomalies. Pathologists integrating their expertise with feedback from an AI system could result in a synergy that outperforms both the individual pathologist and the system. Despite the hype around AI assistance, existing literature on this topic within the pathology domain is limited. We investigated the value of AI assistance for grading prostate biopsies. A panel of 14 observers graded 160 biopsies with and without AI assistance. Using AI, the agreement of the panel with an expert reference standard increased significantly (quadratically weighted Cohen's kappa, 0.799 vs. 0.872; p = 0.019). On an external validation set of 87 cases, the panel showed a significant increase in agreement with a panel of international experts in prostate pathology (quadratically weighted Cohen's kappa, 0.733 vs. 0.786; p = 0.003). In both experiments, on a group-level, AI-assisted pathologists outperformed the unassisted pathologists and the standalone AI system. Our results show the potential of AI systems for Gleason grading, but more importantly, show the benefits of pathologist-AI synergy.

Introduction: Targeted therapy against tumor angiogenesis is widely used in clinical practice for patients with colorectal liver metastases (CRLM). Possible predictive biomarkers for tumor angiogenesis, such as, microvessel density (MVD), hypoxia and cell proliferation, can be determined using immunohistochemical staining. However, patients ineligible for surgical treatment need to undergo invasive diagnostic interventions in order to determine these biomarkers. CT perfusion (CTP) is an emerging functional imaging technique, which can non-invasively determine vascular properties of solid tumors. The purpose of this study was to evaluate CTP with histological biomarkers in CRLM., Material and Methods: Patients with CRLM underwent CTP one day before liver surgery. CTP analysis was performed on the entire volume of the largest metastases in each patient. Dual-input maximum slope analysis was used and data concerning arterial flow (AF), portal flow (PF) and perfusion index (PI) were recorded. Immunohistochemical staining with CD34, M75/CA-IX and MIB-1 was performed on the rim in the midsection of the tumor to determine respectively MVD, hypoxia and cell proliferation., Results: Twenty CRLM in 20 patients were studied. Mean size of the largest CRLM was 37 mm (95% CI 21-54 mm). Mean AF and PF were respectively 64 ml/min/100ml (95% CI 48-79) and 30 ml/min/100ml (95% CI 22-38). Mean PI was 68% (95% CI 62-73). No significant correlation was found between tumor growth patterns and CTP (p = 0.95). MVD did not significantly correlate to AF (r = 0.05; p = 0.84), PF (r = 0.17; p = 0.47) and PI (r = -0.12; p = 0.63). Cell proliferation also did not significantly correlate to AF (r = 0.07; p = 0.78), PF (r = -0.01; p = 0.95) and PI (r = 0.15; p = 0.52). Hypoxia did not significantly correlate to AF (r = -0.05; p = 0.83), however, significantly to PF (r = 0.51; p = 0.02) and a trend to negative correlation with PF (r = -0.43; p = 0.06). However, after controlling the false discovery rate, no significant correlation between CTP and used immunohistochemical biomarkers was found., Conclusion: In conclusion, this feasibility study found a trend to negative correlation between PI and hypoxia, CTP might therefore possibly evaluate this prognostic marker in CRLM non-invasively. However, CTP is not an appropriate technique for the assessment of microvessels or cell proliferation in CRLM., Competing Interests: The authors have declared that no competing interests exist.

Wood and litter degrading fungi are the main decomposers of lignocellulose and thus play a key role in carbon cycling in nature. Here, we provide evidence for a novel lignocellulose degradation strategy employed by the litter degrading fungus Agaricus bisporus (known as the white button mushroom). Fusion of hyphae allows this fungus to synchronize the activity of its mycelium over large distances (50 cm). The synchronized activity has a 13-h interval that increases to 20 h before becoming irregular and it is associated with a 3.5-fold increase in respiration, while compost temperature increases up to 2°C. Transcriptomic analysis of this burst-like phenomenon supports a cyclic degradation of lignin, deconstruction of (hemi-) cellulose and microbial cell wall polymers, and uptake of degradation products during vegetative growth of A. bisporus. Cycling in expression of the ligninolytic system, of enzymes involved in saccharification, and of proteins involved in nutrient uptake is proposed to provide an efficient way for degradation of substrates such as litter., (© 2020 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Background: The literature on the prognostic relevance of signet-ring cell (SRC) histology in gastric cancer (GC) is controversial which is most likely related to inconsistent SRC classification based on haematoxylin-eosin staining. We hypothesised that mucin stains can consistently identify SRC-GC and predict GC patient outcome., Methods: We performed a comprehensive literature review on mucin stains in SRC-GC and characterised the mucin expression in 851 Caucasian GC and 410 Asian GC using Alcian Blue (AB)-Periodic Acid-Schiff (PAS), MUC2 (intestinal-type mucin), and MUC5AC (gastric-type mucin). The relationship between mucin expression and histological phenotype [poorly cohesive (PC) including proportion of SRCs, non-poorly cohesive (non-PC), or mucinous (MC)], clinicopathological variables, and patient outcome was analysed., Results: Depending on mucin expression and cut-offs, the positivity rates of SRC-GC reported in the literature varied from 6 to 100%. Patients with MUC2 positive SRC-GC or SRC-GC with (gastro)intestinal phenotype had poorest outcome. In our cohort study, PC with ≥ 10% SRCs expressed more frequently MUC2, MUC5AC, and ABPAS (p < 0.001, p = 0.004 and p < 0.001, respectively). Caucasians with AB positive GC or combined ABPAS-MUC2 positive and MUC5AC negative had poorest outcome (all p = 0.002). This association was not seen in Asian patients., Conclusions: This is the first study to suggest that mucin stains do not help to differentiate between SRC-GC and non-SRC-GC. However, mucin stains appear to be able to identify GC patients with different outcome. To our surprise, the relationship between outcome and mucin expression seems to differ between Caucasian and Asian GC patients which warrants further investigations.

Agaricus bisporus consumes carbohydrates contained in wheat straw based compost used for commercial mushroom production. Double substituted arabinoxylan is part of the ~40% of the compost polysaccharides that are not degraded by A. bisporus during its growth and development. Genes encoding α-1,3-l-arabinofuranosidase (AXHd3) enzymes that act on xylosyl residues doubly substituted with arabinosyl residues are absent in this mushroom forming fungus. Here, the AXHd3 encoding hgh43 gene of Humicola insolens was expressed in A. bisporus with the aim to improve its substrate utilization and mushroom yield. Transformants secreted active AXHd3 in compost as shown by the degradation of double substituted arabinoxylan oligomers in an in vitro assay. However, carbohydrate composition and degree of arabinosyl substitution of arabinoxylans were not affected in compost possibly due to inaccessibility of the doubly substituted xylosyl residues., Competing Interests: The fact that CNC Grondstoffen B.V. provided compost did neither influence study design, data collection and analysis, decision to publish, writing the manuscript, nor the adherence to PLOS ONE policies on sharing data and materials.

Many fungi are polykaryotic, containing multiple nuclei per cell. In the case of heterokaryons, there are different nuclear types within a single cell. It is unknown what the different nuclear types contribute in terms of mRNA expression levels in fungal heterokaryons. Each cell of the mushroom Agaricus bisporus contains two to 25 nuclei of two nuclear types originating from two parental strains. Using RNA-sequencing data, we assess the differential mRNA contribution of individual nuclear types and its functional impact. We studied differential expression between genes of the two nuclear types, P1 and P2, throughout mushroom development in various tissue types. P1 and P2 produced specific mRNA profiles that changed through mushroom development. Differential regulation occurred at the gene level, rather than at the locus, chromosomal, or nuclear level. P1 dominated mRNA production throughout development, and P2 showed more differentially up-regulated genes in important functional groups. In the vegetative mycelium, P2 up-regulated almost threefold more metabolism genes and carbohydrate active enzymes (cazymes) than P1, suggesting phenotypic differences in growth. We identified widespread transcriptomic variation between the nuclear types of A. bisporus Our method enables studying nucleus-specific expression, which likely influences the phenotype of a fungus in a polykaryotic stage. Our findings have a wider impact to better understand gene regulation in fungi in a heterokaryotic state. This work provides insight into the transcriptomic variation introduced by genomic nuclear separation., Competing Interests: The authors declare no conflict of interest., (Copyright © 2018 the Author(s). Published by PNAS.)

Degradation of lignin by fungi enhances availability of cellulose and hemicellulose in plant waste and thereby increases the amount of carbon source available to these microorganisms. The button mushroom Agaricus bisporus degrades only about half of the lignin in compost and about 40% of the carbohydrates remain unutilized during mushroom cultivation. Here it was assessed whether over-expression of the manganese peroxidase gene mnp1 improves lignin degradation and, as a consequence, carbohydrate breakdown by A. bisporus. Transformants expressing mnp1 under the control of actin regulatory sequences produced MnP activity in malt extract medium, while the parental strain A15 did not. MnP activity was increased 0.3- and 3-fold at casing and after the 2nd flush of a semi-commercial cultivation, respectively, when compared to strain A15. Pyrolysis-GC-MS showed that overexpression of MnP decreased phenylmethane and phenylethane type lignin relative to the phenylpropane type after the 2nd flush. However, it neither affected the syringyl/guaiacyl derived residue ratio nor the ratio of oxidized to non-oxidized lignin residues. Moreover, the carbohydrate content and accessibility was not affected in compost. Notably, the capacity of compost extract to consume the MnP co-factor H 2 O 2 was 4- to 8-fold higher than its production. This may well explain why over-expression of mnp1 did not improve carbohydrate degradation in compost. In fact, availability of H 2 O 2 may limit lignin degradation by wild-type A. bisporus.

Agaricus bisporus mushrooms are commercially produced on a microbe rich compost. Here, fungal and bacterial biomass was quantified in compost with and without colonization by A. bisporus. Chitin content, indicative of total fungal biomass, increased during a 26-day period from 576 to 779 nmol N-acetylglucosamine g -1 compost in the absence of A. bisporus (negative control). A similar increase was found in the presence of this mushroom forming fungus. The fungal phospholipid-derived fatty acid (PLFA) marker C18:2ω6, indicative of the living fraction of the fungal biomass, decreased from 575 to 280 nmol g -1 compost in the negative control. In contrast, it increased to 1200 nmol g -1 compost in the presence of A. bisporus. Laccase activity was absent throughout culturing in the negative control, while it correlated with the fungal PLFA marker in the presence of A. bisporus. PLFA was also used to quantify living bacterial biomass. In the negative control, the bacterial markers remained constant at 3000-3200 nmol PLFA g -1 compost. In contrast, they decreased to 850 nmol g -1 compost during vegetative growth of A. bisporus, implying that bacterial biomass decreased from 17.7 to 4.7 mg g -1 compost. The relative amount of the Gram positive associated PLFA markers a15:0 and a17:0 and the Gram negative PLFA associated markers cy17:0 and cy19:0 increased and decreased, respectively, suggesting that Gram negative bacteria are more suppressed by A. bisporus. Together, these data indicate that fungal biomass can make up 6.8% of the compost after A. bisporus colonization, 57% of which being dead. Moreover, results show that A. bisporus impacts biomass and composition of bacteria in compost.

Malassezia spp. are part of the normal human and animal mycobiota but are also associated with a variety of dermatological diseases. The absence of a transformation system hampered studies to reveal mechanisms underlying the switch from the non-pathogenic to pathogenic life style. Here we describe, a highly efficient Agrobacterium-mediated genetic transformation system for Malassezia furfur and M. pachydermatis. A binary T-DNA vector with the hygromycin B phosphotransferase (hpt) selection marker and the green fluorescent protein gene (gfp) was introduced in M. furfur and M. pachydermatis by combining the transformation protocols of Agaricus bisporus and Cryptococcus neoformans. Optimal temperature and co-cultivation time for transformation were 5 and 7days at 19°C and 24°C, respectively. Transformation efficiency was 0.75-1.5% for M. furfur and 0.6-7.5% for M. pachydermatis. Integration of the hpt resistance cassette and gfp was verified using PCR and fluorescence microscopy, respectively. The T-DNA was mitotically stable in approximately 80% of the transformants after 10 times sub-culturing in the absence of hygromycin. Improving transformation protocols contribute to study the biology and pathophysiology of Malassezia., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose: To determine the full spectrum of ocular manifestations in patients with Noonan syndrome (NS)., Design: Prospective cross-sectional clinical and genetic study in a tertiary referral center., Participants: Twenty-five patients with NS (mean age, 14 years; range, 8 months-25 years) clinically diagnosed by validated criteria., Methods: All patients were examined by the same team following a detailed study protocol. Genetic analyses were performed in 23 patients., Main Outcome Measures: Ocular abnormalities of vision and refraction, external ocular features, ocular position and motility, anterior segment, posterior segment, and intraocular pressure., Results: Ocular features of vision and refraction were amblyopia (32%), myopia (40%), and astigmatism (52%). External ocular features were epicanthic folds (84%), hypertelorism (68%), ptosis (56%), high upper eyelid crease (64%), lower eyelid retraction (60%), abnormal upward slanting palpebral fissures (36%), downward slanting palpebral fissures (32%), and lagophthalmos (28%). Orthoptic abnormalities included strabismus (40%), abnormal stereopsis (44%), and limited ocular motility (40%). Anterior segment abnormalities included prominent corneal nerves (72%) and posterior embryotoxon (32%). Additional ocular features were found, including nonglaucomatous optic disc excavation (20%), relatively low (<10 mmHg) intraocular pressure (22%), and optic nerve hypoplasia (4%). Mutations were established in 22 patients: 19 PTPN11 mutations (76%), 1 SOS1 mutation, 1 BRAF mutation, and 1 KRAS mutation. The patient with the highest number of prominent corneal nerves had an SOS1 mutation. The patient with the lowest visual acuity, associated with bilateral optic nerve hypoplasia, had a BRAF mutation. Patients with severe ptosis and nearly total absence of levator muscle function had PTPN11 mutations. All patients showed at least 3 ocular features (range, 3-13; mean, 7), including at least 1 external ocular feature in more than 95% of the patients., Conclusions: Noonan syndrome is a clinical diagnosis with multiple genetic bases associated with an extensive variety of congenital ocular abnormalities. Ocular features of NS are characterized by 1 or more developmental anomalies of the eyelids (involving the position, opening, and closure) associated with various other ocular abnormalities in childhood, including amblyopia, myopia, astigmatism, strabismus, limited ocular motility, prominent corneal nerves, and posterior embryotoxon., (Copyright © 2016 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

The Cys2His2 zinc finger protein gene c2h2 of Schizophyllum commune is involved in mushroom formation. Its inactivation results in a strain that is arrested at the stage of aggregate formation. In this study, the c2h2 orthologue of Agaricus bisporus was over-expressed in this white button mushroom forming basidiomycete using Agrobacterium-mediated transformation. Morphology, cap expansion rate, and total number and biomass of mushrooms were not affected by over-expression of c2h2. However, yield per day of the c2h2 over-expression strains peaked 1 day earlier. These data and expression analysis indicate that C2H2 impacts timing of mushroom formation at an early stage of development, making its encoding gene a target for breeding of commercial mushroom strains.

The α7 nicotinic acetylcholine receptor (nAChR) belongs to the family of pentameric ligand-gated ion channels and is involved in fast synaptic signaling. In this study, we take advantage of a recently identified chimera of the extracellular domain of the native α7 nicotinic acetylcholine receptor and acetylcholine binding protein, termed α7-AChBP. This chimeric receptor was used to conduct an innovative fragment-library screening in combination with X-ray crystallography to identify allosteric binding sites. One allosteric site is surface-exposed and is located near the N-terminal α-helix of the extracellular domain. Ligand binding at this site causes a conformational change of the α-helix as the fragment wedges between the α-helix and a loop homologous to the main immunogenic region of the muscle α1 subunit. A second site is located in the vestibule of the receptor, in a preexisting intrasubunit pocket opposite the agonist binding site and corresponds to a previously identified site involved in positive allosteric modulation of the bacterial homolog ELIC. A third site is located at a pocket right below the agonist binding site. Using electrophysiological recordings on the human α7 nAChR we demonstrate that the identified fragments, which bind at these sites, can modulate receptor activation. This work presents a structural framework for different allosteric binding sites in the α7 nAChR and paves the way for future development of novel allosteric modulators with therapeutic potential.

We present the case of an 82-year-old woman undergoing high-risk chronic total occlusion percutaneous coronary intervention (CTO-PCI) of the right coronary artery. Hours after the procedure, a subepicardial haematoma was diagnosed as a result of coronary perforation during the procedure. This rare and potentially lethal complication evolved exceptionally benignly after conservative management; our patient fully recovered. Increasingly complex procedures in high-risk patient categories warrant awareness of procedural complications, especially those that are subtle and appear relatively late, and are therefore most hazardous. Recognition of this rare complication and choosing the optimal strategy is of the utmost importance when dealing with patients who undergo PCI. We here describe the rare case of a potential lethal complication in high-risk CTO-PCI, which evolved relatively benignly., (2014 BMJ Publishing Group Ltd.)

Cardiac pulsatility and aortic compliance may result in aortic area and diameter changes throughout the cardiac cycle in the entire aorta. Until this moment these dynamic changes could never be established in the aortic root (aortic annulus, sinuses of Valsalva and sinotubular junction). The aim of this study was to visualize and characterize the changes in aortic root dimensions during systole and diastole with ECG-gated multidetector row computed tomography (MDCT). MDCT scans of subjects without aortic root disease were analyzed. Retrospectively, ECG-gated reconstructions at each 10% of the cardiac cycle were made and analyzed during systole (30-40%) and diastole (70-75%). Axial planes were reconstructed at three different levels of the aortic root. At each level the maximal and its perpendicular luminal dimension were measured. The mean dimensions of the total study group (n = 108, mean age 56 ± 13 years) do not show any significant difference between systole and diastole. The individual dimensions vary up to 5 mm. However, the differences range between minus 5 mm (diastolic dimension is greater than systolic dimensions) and 5 mm (vice versa). This variability is independent of gender, age, height and weight. This study demonstrated a significant individual dynamic change in the dimensions of the aortic root. These results are highly unpredictable. Most of the healthy subjects have larger systolic dimensions, however, some do have larger diastolic dimensions.

Realistic representation of protein flexibility in biomolecular simulations remains an unsolved fundamental problem and is an active area of research. The high flexibility of the cytochrome P450 2D6 (CYP2D6) active site represents a challenge for accurate prediction of the preferred binding mode and site of metabolism (SOM) for compounds metabolized by this important enzyme. To account for this flexibility, we generated a large ensemble of unbiased CYP2D6 conformations, to which small molecule substrates were docked to predict their experimentally observed SOM. SOM predictivity was investigated as a function of the number of protein structures, the scoring function, the SOM-heme cutoff distance used to distinguish metabolic sites, and intrinsic reactivity. Good SOM predictions for CYP2D6 require information from the protein. A critical parameter is the distance between the heme iron and the candidate site of metabolism. The best predictions were achieved with cutoff distances consistent with the chemistry relevant to CYP2D6 metabolism. Combination of the new ensemble-based docking method with estimated intrinsic reactivities of substrate sites considerably improved the predictivity of the model. Testing on an independent set of substrates yielded area under curve values as high as 0.93, validating our new approach.

The development of HIV integrase (IN) strand transfer inhibitors (INSTIs) and our understanding of viral resistance to these molecules have been hampered by a paucity of available structural data. We recently reported cocrystal structures of the prototype foamy virus (PFV) intasome with raltegravir and elvitegravir, establishing the general INSTI binding mode. We now present an expanded set of cocrystal structures containing PFV intasomes complexed with first- and second-generation INSTIs at resolutions of up to 2.5 Å. Importantly, the improved resolution allowed us to refine the complete coordination spheres of the catalytic metal cations within the INSTI-bound intasome active site. We show that like the Q148H/G140S and N155H HIV-1 IN variants, the analogous S217H and N224H PFV INs display reduced sensitivity to raltegravir in vitro. Crystal structures of the mutant PFV intasomes in INSTI-free and -bound forms revealed that the amino acid substitutions necessitate considerable conformational rearrangements within the IN active site to accommodate an INSTI, thus explaining their adverse effects on raltegravir antiviral activity. Furthermore, our structures predict physical proximity and an interaction between HIV-1 IN mutant residues His148 and Ser/Ala140, rationalizing the coevolution of Q148H and G140S/A mutations in drug-resistant viral strains.

The RNA-dependent RNA polymerase (NS5B) of hepatitis C virus (HCV) is an unusually attractive target for drug discovery since it contains five distinct drugable sites. The success of novel antiviral therapies will require nonnucleoside inhibitors to be active in at least patients infected with HCV of subtypes 1a and 1b. Therefore, the genotypic assessment of these agents against clinical isolates derived from genotype 1-infected patients is an important prerequisite for the selection of suitable candidates for clinical development. Here we report the 1a/1b subtype profiling of polymerase inhibitors that bind at each of the four known nonnucleoside binding sites. We show that inhibition of all of the clinical isolates tested is maintained, except for inhibitors that bind at the palm-1 binding site. Subtype coverage varies across chemotypes within this class of inhibitors, and inhibition of genotype 1a improves when hydrophobic contact with the polymerase is increased. We investigated if the polymorphism of the palm-1 binding site is the sole cause of the reduced susceptibility of subtype 1a to inhibition by 1,5-benzodiazepines by using reverse genetics, X-ray crystallography, and surface plasmon resonance studies. We showed Y415F to be a key determinant in conferring resistance on subtype 1a, with this effect being mediated through an inhibitor- and enzyme-bound water molecule. Binding studies revealed that the mechanism of subtype 1a resistance is faster dissociation of the inhibitor from the enzyme.

Objectives: This study sought to determine the diagnostic accuracy of 64-slice computed tomographic coronary angiography (CTCA) to detect or rule out significant coronary artery disease (CAD)., Background: CTCA is emerging as a noninvasive technique to detect coronary atherosclerosis., Methods: We conducted a prospective, multicenter, multivendor study involving 360 symptomatic patients with acute and stable anginal syndromes who were between 50 and 70 years of age and were referred for diagnostic conventional coronary angiography (CCA) from September 2004 through June 2006. All patients underwent a nonenhanced calcium scan and a CTCA, which was compared with CCA. No patients or segments were excluded because of impaired image quality attributable to either coronary motion or calcifications. Patient-, vessel-, and segment-based sensitivities and specificities were calculated to detect or rule out significant CAD, defined as >or=50% lumen diameter reduction., Results: The prevalence among patients of having at least 1 significant stenosis was 68%. In a patient-based analysis, the sensitivity for detecting patients with significant CAD was 99% (95% confidence interval [CI]: 98% to 100%), specificity was 64% (95% CI: 55% to 73%), positive predictive value was 86% (95% CI: 82% to 90%), and negative predictive value was 97% (95% CI: 94% to 100%). In a segment-based analysis, the sensitivity was 88% (95% CI: 85% to 91%), specificity was 90% (95% CI: 89% to 92%), positive predictive value was 47% (95% CI: 44% to 51%), and negative predictive value was 99% (95% CI: 98% to 99%)., Conclusions: Among patients in whom a decision had already been made to obtain CCA, 64-slice CTCA was reliable for ruling out significant CAD in patients with stable and unstable anginal syndromes. A positive 64-slice CTCA scan often overestimates the severity of atherosclerotic obstructions and requires further testing to guide patient management.

Fat surrounding coronary arteries might aggravate coronary artery disease (CAD). We investigated the relation between epicardial adipose tissue (EAT) and pericoronary fat and coronary atherosclerosis and coronary artery calcium (CAC) in patients with suspected CAD and whether this relation is modified by total body weight. This was a cross-sectional study of 128 patients with angina pectoris (61 +/- 6 years of age) undergoing coronary angiography. EAT volume and pericoronary fat thickness were measured with cardiac computed tomography. Severity of coronary atherosclerosis was assessed by the number of stenotic (> or =50%) coronary vessels; extent of CAC was determined by the Agatston score. Patients were stratified for median total body weight (body mass index [BMI] 27 kg/m(2)). Overall, EAT and pericoronary fat were not associated with severity of coronary atherosclerosis and extent of CAC. In patients with low BMI, those with multivessel disease had increased EAT volume (100 vs 67 cm(3), p = 0.04) and pericoronary fat thickness (9.8 vs 8.4 mm, p = 0.06) compared with those without CAD. Also, patients with severe CAC had increased EAT volume (108.0 vs 69 cm(3), p = 0.02) and pericoronary fat thickness (10.0 vs 8.2 mm, p value = 0.01) compared with those with minimal/absent CAC. In conclusion, EAT and pericoronary fat were not associated with severity of coronary atherosclerosis and CAC in patients with suspected CAD. However, in those with low BMI, increased EAT and pericoronary fat were related to more severe coronary atherosclerosis and CAC. Fat surrounding coronary arteries may be involved in the process of coronary atherosclerosis, although this is different for patients with low and high BMIs.

Background: Peripheral arterial disease (PAD) is a common disease associated with a considerably increased risk of future cardiovascular events and most of these patients will die from coronary artery disease (CAD). Screening for silent CAD has become an option with recent non-invasive developments in CT (computed tomography)-angiography and MR (magnetic resonance) stress testing. Screening in combination with more aggressive treatment may improve prognosis. Therefore we propose to study whether a cardiac imaging algorithm, using non-invasive imaging techniques followed by treatment will reduce the risk of cardiovascular disease in PAD patients free from cardiac symptoms., Design: The GROUND study is designed as a prospective, multi-center, randomized clinical trial. Patients with peripheral arterial disease, but without symptomatic cardiac disease will be asked to participate. All patients receive a proper risk factor management before randomization. Half of the recruited patients will enter the 'control group' and only undergo CT calcium scoring. The other half of the recruited patients (index group) will undergo the non invasive cardiac imaging algorithm followed by evidence-based treatment. First, patients are submitted to CT calcium scoring and CT angiography. Patients with a left main (or equivalent) coronary artery stenosis of > 50% on CT will be referred to a cardiologist without further imaging. All other patients in this group will undergo dobutamine stress magnetic resonance (DSMR) testing. Patients with a DSMR positive for ischemia will also be referred to a cardiologist. These patients are candidates for conventional coronary angiography and cardiac interventions (coronary artery bypass grafting (CABG) or percutaneous cardiac interventions (PCI)), if indicated. All participants of the trial will enter a 5 year follow up period for the occurrence of cardiovascular events. Sequential interim analysis will take place. Based on sample size calculations about 1200 patients are needed to detect a 24% reduction in primary outcome., Implications: The GROUND study will provide insight into the question whether non-invasive cardiac imaging reduces the risk of cardiovascular events in patients with peripheral arterial disease, but without symptoms of coronary artery disease., Trial Registration: Clinicaltrials.gov NCT00189111.

Objective: Adipose tissue surrounding coronary arteries may contribute to the development of coronary atherosclerosis given its localisation and potential for local production of inflammatory cytokines. We compared various measurements for quantifying epicardial adipose tissue (EAT) and peri-coronary fat using cardiac CT. Additionally, we estimated their relationship with obesity and metabolic syndrome in patients suspected of coronary artery disease (CAD)., Methods: EAT and peri-coronary fat measurements were performed on cardiac multi-slice CT scans in 60 patients (aged 50-70 years) referred for coronary angiography. EAT was measured as thickness on the right ventricular free wall, as area at the base of the ventricles, and as volume. Peri-coronary fat was assessed as thickness and cross-sectional area surrounding the three main coronary arteries. Linear regression analysis was used to assess the relation of EAT and peri-coronary fat with obesity and metabolic syndrome (ATPIII criteria)., Results: Volumetric EAT measurements showed good reproducibility with low coefficients of variation (CVs) varying between 3.0% and 5.0%. Measurements of EAT and peri-coronary fat thickness and area were moderately reproducible (CVs 11.0-23.4%). The amount of EAT and peri-coronary fat (per standard deviation) was related with obesity (BMI > or =30 kg/m(2)) (beta 1.24; 95% CI 0.66; 1.81) and metabolic syndrome (beta 0.81; 95% CI 0.28; 1.33)., Conclusions: Volumetric quantification of EAT using cardiac CT is highly reproducible compared to more simple measurements as EAT and peri-coronary fat thickness and area. The quantity of EAT and peri-coronary fat is related with the presence of obesity and metabolic syndrome in patients suspected of CAD.

Aims: To determine whether peri-coronary epicardial adipose tissue (EAT) is associated with vascular risk factors and coronary atherosclerosis., Methods and Results: In this study, 573 healthy post-menopausal women underwent a cardiac CT scan to assess coronary calcification. Peri-coronary EAT thickness was measured in the areas of right coronary artery (RCA), left anterior descending (LAD) artery, and left circumflex (LCX) coronary artery. Average EAT thickness was 16.5 +/- 4.3 mm (range 5.9-34.6) in the RCA area, 6.4 +/- 2.2 mm (range 2.0-14.0) in the LAD area, and 10.8 +/- 3.0 mm (range 2.8-29.1) in the LCX area. Overall average thickness was 11.2 +/- 2.2 mm (range 5.4-19.1). EAT was positively related to age (P = 0.002). In age-adjusted linear regression models, EAT was positively related to weight (P< 0.001), waist circumference (P< 0.001), waist-to-hip ratio (P< 0.001), body mass index (P< 0.001), glucose (P< 0.001), triglycerides (P = 0.001), use of anti-hypertensive drugs (P = 0.007), and systolic blood pressure (P = 0.034), and inversely to HDL cholesterol (P = 0.005). In multivariable models, age, weight, waist circumference, smoking, and glucose were the main determinants of EAT. EAT showed a graded relation with coronary calcification (P = 0.026)., Conclusion: EAT is strongly related to vascular risk factors and coronary calcification. Our findings support the hypothesis that EAT affects coronary atherosclerosis and possibly coronary risk.

Objective: To study how much the calcium scores at various phases throughout the cardiac cycle deviate from the score in the most motionless phase during retrospectively electrocardiogram (ECG)-gated multidetector row computed tomography (MDCT) of the heart and to evaluate how to optimize ECG-based tube current modulation so that errors in calcium scoring can be minimized while dose savings can be maximized., Materials and Methods: In 73 subjects with known or suspected coronary artery disease we performed retrospectively ECG-gated 64-detector row computed tomography for calcium scoring. Four subjects were excluded after scanning because of breathing artifacts or lack of coronary calcification. The scans of 69 subjects (46 men, mean age 62 +/- 6 years) were used for further analysis. Heart rate during the scan was recorded. In each patient, calcium scoring [Agatston score (AS), mass score (MS), and volume score, (VS)] was performed on 10 data sets reconstructed at 10%-intervals throughout the cardiac cycle. The most motionless phase was subjectively determined and used as the reference phase. For the score in each phase, deviation from the score in the reference phase was determined. An ECG-simulator was used to determine the amount of dose saving while scanning with dose modulation and applying diagnostic dose during 1 or several phases., Results: Mean heart rate was 63 (+/-13) beats per minute (bpm). In 51% of patients the reference phase was the 70% phase. Using the calcium score in the 70% phase (mid-diastole) instead of the reference at heart rates below 70 bpm would have induced a median score deviation of 0% [interquartile range: 0%-6% (AS, MS, and VS)] and using the calcium score in the 40% phase (end-systole) at heart rates > or =70 bpm would also have induced a median score deviation of 0% [interquartile range: 0%-7% (AS), 0%-5% (MS), and 0%-3% (VS)]. Errors in calcium scores of more than 10% occur in around 10% of subjects for all 3 scoring algorithms. Dose savings increased with lower heart rates and shorter application of diagnostic dose., Conclusions: The optimum phases for dose modulation are 70% (mid-diastole) at heart rates below 70 bpm and 40% (end-systole) at heart rates above 70 bpm. Under these conditions dose saving is maximum and a median error of 0% is found for the various calcium scoring techniques with score errors of more than 10% in around 10% of subjects.

Lamotrigine (LTG, Lamictal), one of the newer antiepileptic drugs, was admitted to the Dutch market in 1996. It was first used as adjunctive therapy and later as a monotherapy in partial and generalized epilepsy. All patients who started on LTG in 1996 or 1997 in the Epilepsy Centre Kempenhaeghe (n=314) were enrolled in this study and followed for 48 months. The data indicate that the retention rates for LTG after 1, 2, 3, and 4 years are respectively 74.4, 69.3, 63.1, and 55.6%. Patients with normal cognitive function were more likely to continue than patients with mental retardation. The main reason for discontinuing LTG therapy was lack of efficacy (19.1%). Four patients (1.4%) were seizure-free for the total follow-up period of 48 months. The most frequently reported negative side effects were dizziness and headache, both in patients who continued and in those who discontinued therapy. A large percentage of patients also reported positive side effects like "feeling/being more active" and "feeling more clear/more responsive." For the whole patient group, the plasma level of LTG was measured 277 times. Plasma levels of LTG were influenced by the patients' comedications. Plasma levels of LTG in groups taking LTG in monotherapy, LTG plus an inducer, and LTG plus valproate were 8.7, 4.8, and 8.7 mg/L, respectively. The correlation between measured plasma level and dose confirm the manufacturer's dose recommendations. The manufacturer recommends half the dosage of lamotrigine monotherapy when the patient also uses valproate. When the patient uses an inducer, the dosage of LTG must be two times the dose used in monotherapy.

Background: Ptosis and dysphagia are important features in oculopharyngeal muscular dystrophy (OPMD)., Objective: Retroflexion of the head is a well known compensatory mechanism for ptosis, but generally retroflexion has a negative effect on swallowing. We hypothesised that severity of ptosis is related to degree of retroflexion and that this compensation is responsible for deteriorating dysphagia., Methods: Nine OPMD patients were examined in the conditions "head position adapted to ptosis" and "head position slightly flexed". Ptosis was quantified by photogrammetry and retroflexion of the head by digital photographs. The severity of dysphagia was measured using visual analogue scales (VAS) and by calculating swallowing volumes and oropharyngeal swallow efficiency (OPSE) based on videofluoroscopy., Results: Statistical analyses show a significant relationship between ptosis and degree of retroflexion. The degree of retroflexion of the head correlated significantly with VAS scores and with the maximum swallowing volume. The slightly flexed head position significantly improved VAS scores as well as swallowing volumes and OPSE., Conclusion: In OPMD patients, ptosis significantly correlates with retroflexion of the head, which has a negative effect on swallowing. Subjective and objective reduction of swallowing problems was found when patients were instructed to eat and drink with a slightly flexed head position.

Conventional coronary angiography (CAG) has been the reference standard for the assessment of coronary artery disease since its introduction in 1958. However, several studies have shown that diagnostic CAG has an average morbidity of 2% and a mortality of approximately 0.1%. In the last decade, progress in medical imaging has opened the way to noninvasive assessment of the coronary arteries at lower cost and risk. Of the different modalities, multislice CT (MSCT) has made the biggest step forward. At the 2005 European Congress of Radiology (ECR), experiences with the latest developments in noninvasive coronary artery imaging were reported. This report summarises the advances in the use of MSCT in coronary stenosis detection, emergency decision-making, plaque imaging, and the analysis of cardiac function and late enhancement. Also, attention is paid to new strategies to reduce MSCT-related radiation exposure.

Here we report unexpected findings in a 17-year-old female patient referred for coronary angiography and percutaneous intervention. During the angiography we observed a complete occlusion of the left main coronary artery (LMCA). The occurrence of coronary abnormalities at this age is extremely rare and mostly caused by congenital abnormalities. The diagnosis of premature atherosclerosis at this age is unlikely unless the patient suffers from severe lipoprotein disease. Here we describe a rare case of LMCA occlusion, the most likely cause of the disease and the potential implications for therapy.