Combining fracture mechanics and rheology to investigate the impact of micro-aeration on chocolate oral processing.

This study presents a rigorous mechanical characterisation investigation on milk chocolate with varying porosities, at different temperatures and strain rate levels. Uniaxial compression tests at temperatures varying from 20 °C to 30 °C were performed to measure the bulk properties of chocolate as a function of porosity and temperature. Fracture experiments were also conducted to compute the fracture energy at temperature levels between 20 °C and 30 °C for all tested samples. Additionally, rheological experiments are conducted to compute the viscosity of the different chocolates at 37 °C. This combined experimental analysis of solid mechanics, fracture mechanics, and rheology aims to define the impact of temperature and chocolate's phase change from solid to liquid on its mechanical properties. Moreover, the impact of micro-aeration on the relationship between material properties and temperature is discussed. The results demonstrate a significant impact of both temperature and micro-aeration on the chocolate's material properties; fracture stresses decrease with micro-aeration due to the presence of micro-pores creating weak links in the chocolate matrix, the critical strain energy release rate decreases with micro-aeration at temperatures up to 25 °C and increases at temperatures above 30 °C. Finally, the viscosity at 37 °C increases with increasing porosity due to the obstruction of the flow by micro-pores acting as "solid" particles. The results highlight how the impact of micro-aeration on the material properties of chocolate alters as the testing temperature rises and the material changes phase. The relationships between the micro-aeration and material properties and the dependence of temperature on the different mechanical properties are used to explain the difference in textural attributes as obtained from temporal dominance sensation tests. This study seeks to contribute valuable insights into the field of chocolate technology, emphasizing the need for a combined engineering approach to understand the structural breakdown of chocolate during oral processing as mechanisms such as chewing, melting, mixing and shearing occur.

[Postoperative management following decompressive hemicraniectomy for malignant middle cerebral artery infarction-A German nationwide survey study]. / Postoperatives Management nach dekompressiver Hemikraniektomie bei malignem Mediainfarkt ­ eine deutschlandweite Umfragestudie.

BACKGROUND: Malignant middle cerebral artery infarction is a potentially life-threatening disease. Decompressive hemicraniectomy constitutes an evidence-based treatment practice, especially in patients under 60 years of age; however, recommendations with respect to postoperative management and particularly duration of postoperative sedation lack standardization. OBJECTIVE: This survey study aimed to analyze the current situation of patients with malignant middle cerebral artery infarction following hemicraniectomy in the neurointensive care setting. MATERIAL AND METHODS: From 20 September 2021 to 31 October 2021, 43 members of the initiative of German neurointensive trial engagement (IGNITE) network were invited to participate in a standardized anonymous online survey. Descriptive data analysis was performed. RESULTS: Out of 43 centers 29 (67.4%) participated in the survey, including 24 university hospitals. Of the hospitals 21 have their own neurological intensive care unit. While 23.1% favored a standardized approach regarding postoperative sedation, the majority utilized individual criteria (e.g., intracranial pressure increase, weaning parameters, complications) to assess the need and duration. The timing of targeted extubation varied widely between hospitals (≤ 24 h 19.2%, ≤ 3 days in 30.8%, ≤ 5 days in 19.2%, > 5 days in 15.4%). Early tracheotomy (≤ 7 days) is performed in 19.2% and 80.8% of the centers aim for tracheotomy within 14 days. Hyperosmolar treatment is used on a regular basis in 53.9% and 22 centers (84.6%) agreed to participate in a clinical trial addressing the duration of postoperative sedation and ventilation. CONCLUSION: The results of this nationwide survey among neurointensive care units in Germany reflect a remarkable heterogeneity in the treatment practices of patients with malignant middle cerebral artery infarction undergoing hemicraniectomy, especially with respect to the duration of postoperative sedation and ventilation. A randomized trial in this matter seems warranted.

Chemical and electronic structure of the heavily intermixed (Cd,Zn)S:Ga/CuSbS2 interface.

The interface formation and chemical and electronic structure of the (Cd,Zn)S:Ga/CuSbS2 thin-film solar cell heterojunction were studied using hard X-ray photoelectron spectroscopy (HAXPES) of the bare absorber and a buffer/absorber sample set for which the buffer thickness was varied between 1 and 50 nm. We find a heavily intermixed interface, involving Cu, Zn, and Cd as well as significant Ga and Cu profiles in the buffer. The valence band (VB) offset at the buffer/absorber interface was derived as (-1.3 ± 0.1) eV, which must be considered an upper bound as the Cu diffused into the buffer might form a Cu-derived VB maximum located closer to the Fermi level. The estimated conduction band minimum was 'cliff'-like; a situation made more severe considering the Cu-deficiency found for the CuSbS2 surface. The complex interface structure's effect on the performance of (Cd,Zn)S:Ga/CuSbS2-based solar cells and its limitation is discussed together with possible mitigation strategies.

Experimental and numerical evaluation of the effect of micro-aeration on the thermal properties of chocolate.

Thermal properties, such as thermal conductivity, specific heat capacity and latent heat, influence the melting and solidification of chocolate. The accurate prediction of these properties for micro-aerated chocolate products with varying levels of porosity ranging from 0% to 15% is beneficial for understanding and control of heat transfer mechanisms during chocolate manufacturing and food oral processing. The former process is important for the final quality of chocolate and the latter is associated with sensorial attributes, such as grittiness, melting time and flavour. This study proposes a novel multiscale finite element model to accurately predict the temporal and spatial evolution of temperature across chocolate samples. The model is evaluated via heat transfer experiments at temperatures varying from 16 °C to 45 °C. Both experimental and numerical results suggest that the rate of heat transfer within the micro-aerated chocolate is reduced by 7% when the 15% micro-aerated chocolate is compared to its solid counterpart. More specifically, on average, the thermal conductivity decreased by 20% and specific heat capacity increased by 10% for 15% micro-aeration, suggesting that micro-pores act as thermal barriers to heat flow. The latter trend is unexpected for porous materials and thus the presence of a third phase at the pore's interface is proposed which might store thermal energy leading to a delayed release to the chocolate system. The developed multiscale numerical model provides a design tool to create pore structures in chocolate with optimum melting or solidifying response.

Seasonal shifts in isoprenoid emission composition from three hyperdominant tree species in central Amazonia.

Volatile isoprenoids regulate plant performance and atmospheric processes, and Amazon forests comprise the dominant source to the global atmosphere. Still, there is a poor understanding of how isoprenoid emission capacities vary in response to ecophysiological and environmental controls in Amazonian ecosystems. We measured isoprenoid emission capacities of three Amazonian hyperdominant tree species - Protium hebetatum, Eschweilera grandiflora, Eschweilera coriacea - across seasons and along a topographic and edaphic environmental gradient in the central Amazon. From wet to dry season, both photosynthesis and isoprene emission capacities strongly declined, while emissions increased among the heavier isoprenoids: monoterpenes and sesquiterpenes. Plasticity across habitats was most evident in P. hebetatum, which emitted sesquiterpenes only in the dry season, at rates that significantly increased along the hydro-topographic gradient from white sands (shallow root water access) to uplands (deep water table). We suggest that emission composition shifts are part of a plastic response to increasing abiotic stress (e.g. heat and drought) and reduced photosynthetic supply of substrates for isoprenoid synthesis. Our comprehensive measurements suggest that more emphasis should be placed on other isoprenoids, besides isoprene, in the context of abiotic stress responses. Shifting emission compositions have implications for atmospheric responses because of the strong variation in reactivity among isoprenoid compounds.

Effect of micro-aeration on the mechanical behaviour of chocolates and implications for oral processing.

Aeration in foods has been widely utilised in the food industry to develop novel foods with enhanced sensorial characteristics. Specifically, aeration at the micron-sized scale has a significant impact on the microstructure where micro-bubbles interact with the other microstructural features in chocolates. This study aims to determine the effect of micro-aeration on the mechanical properties of chocolate products, which are directly correlated with textural attributes such as hardness and crumbliness. Uniaxial compression tests were performed to determine the mechanical properties such as Poisson's ratio, Young's modulus and macroscopic yield strength together with fracture tests to estimate the fracture toughness. In vivo mastication tests were also conducted to investigate the link between the fracture properties and fragmentation during the first two chewing cycles. The uniaxial stress-strain data were used to calibrate a viscoplastic constitutive law. The results showed that micro-aeration significantly affects mechanical properties such as Young's modulus, yield and fracture stresses, as well as fracture toughness. In addition, it enhances the brittle nature of the chocolate, as evidenced by lower fracture stress but also lower fracture toughness leading to higher fragmentation, in agreement with observations in the in vivo mastication tests. As evidenced by the XRT images and the stress-strain measurements micro-aeration hinders the re-arrangement of the microscopic features inside the chocolate during the material's deformation. The work provides a new insight of the role of bubbles on the bulk behaviour of complex multiphase materials, such as chocolates, and defines the mechanical properties which are important input parameters for the development of oral processing simulations.

Diffuse glioneuronal tumour with oligodendroglioma-like features and nuclear clusters (DGONC) - a molecularly defined glioneuronal CNS tumour class displaying recurrent monosomy 14.

AIMS: DNA methylation-based central nervous system (CNS) tumour classification has identified numerous molecularly distinct tumour types, and clinically relevant subgroups among known CNS tumour entities that were previously thought to represent homogeneous diseases. Our study aimed at characterizing a novel, molecularly defined variant of glioneuronal CNS tumour. PATIENTS AND METHODS: DNA methylation profiling was performed using the Infinium MethylationEPIC or 450 k BeadChip arrays (Illumina) and analysed using the 'conumee' package in R computing environment. Additional gene panel sequencing was also performed. Tumour samples were collected at the German Cancer Research Centre (DKFZ) and provided by multinational collaborators. Histological sections were also collected and independently reviewed. RESULTS: Genome-wide DNA methylation data from >25 000 CNS tumours were screened for clusters separated from established DNA methylation classes, revealing a novel group comprising 31 tumours, mainly found in paediatric patients. This DNA methylation-defined variant of low-grade CNS tumours with glioneuronal differentiation displays recurrent monosomy 14, nuclear clusters within a morphology that is otherwise reminiscent of oligodendroglioma and other established entities with clear cell histology, and a lack of genetic alterations commonly observed in other (paediatric) glioneuronal entities. CONCLUSIONS: DNA methylation-based tumour classification is an objective method of assessing tumour origins, which may aid in diagnosis, especially for atypical cases. With increasing sample size, methylation analysis allows for the identification of rare, putative new tumour entities, which are currently not recognized by the WHO classification. Our study revealed the existence of a DNA methylation-defined class of low-grade glioneuronal tumours with recurrent monosomy 14, oligodendroglioma-like features and nuclear clusters.

Distinct cortical responses evoked by electrical stimulation of the thalamic ventral intermediate nucleus and of the subthalamic nucleus.

OBJECTIVE: To investigate the spatial and temporal pattern of cortical responses evoked by deep brain stimulation (DBS) of the subthalamic nucleus (STN) and ventral intermediate nucleus of the thalamus (VIM). METHODS: We investigated 7 patients suffering from Essential tremor (ET) and 7 patients with Parkinson's Disease (PD) following the implantation of DBS electrodes (VIM for ET patients, STN for PD patients). Magnetoencephalography (MEG) was used to record cortical responses evoked by electric stimuli that were applied via the DBS electrode in trains of 5 Hz. Dipole fitting was applied to reconstruct the origin of evoked responses. RESULTS: Both VIM and STN DBS led to short latency cortical responses at about 1 ms. The pattern of medium and long latency cortical responses following VIM DBS consisted of peaks at 13, 40, 77, and 116 ms. The associated equivalent dipoles were localized within the central sulcus, 3 patients showed an additional response in the cerebellum at 56 ms. STN DBS evoked cortical responses peaking at 4 ms, 11 ms, and 27 ms, respectively. While most dipoles were localized in the pre- or postcentral gyrus, the distribution was less homogenous compared to VIM stimulation and partially included prefrontal brain areas. CONCLUSION: MEG enables localization of cortical responses evoked by DBS of the VIM and the STN, especially in the sensorimotor cortex. Short latency responses of 1 ms suggest cortical modulation which bypasses synaptic transmission, i.e. antidromic activation of corticofugal fiber pathways.

Rare compound heterozygous variants in PNKP identified by whole exome sequencing in a German patient with ataxia-oculomotor apraxia 4 and pilocytic astrocytoma.

Ataxia-oculomotor apraxia type 4 (AOA4) is a rare autosomal recessive neurologic disorder. The phenotype is characterized by ataxia, oculomotor apraxia, peripheral neuropathy and dystonia. AOA4 is caused by biallelic pathogenic variants in the PNKP gene encoding a polynucleotide kinase 3'-phosphatase with an important function in DNA-damage repair. By whole exome sequencing, we identified 2 variants within the PNKP gene in a 27-year-old German woman with a clinical AOA phenotype combined with a cerebellar pilocytic astrocytoma diagnosed at 23 years of age. One variant, a duplication in exon 14 resulting in the frameshift c.1253_1269dup p.(Thr424fs*49), has previously been described as pathogenic, for example, in cases of AOA4. The second variant, representing a nonsense mutation in exon 17, c.1545C>G p.(Tyr515*), has not yet been described and is predicted to cause a loss of the 7 C-terminal amino acids. This is the first description of AOA4 in a patient with central European descent. Furthermore, the occurrence of a pilocytic astrocytoma has not been described before in an AOA4 patient. Our data demonstrate compound heterozygous PNKP germline variants in a German patient with AOA4 and provide evidence for a possible link with tumor predisposition. Localization of the 2 variants in human PNKP NP_009185.2. NM_007254.3:c.1253_1269dup p.(Thr424fs*49) is predicted to cause a frameshift within the kinase domain, NM_007254.3:c.1545C>G p.(Tyr515*) is predicted to cause loss of 2 C-terminal amino acids of the kinase domain and 5 additional C-terminal amino acids.

Girsanov reweighting for path ensembles and Markov state models.

The sensitivity of molecular dynamics on changes in the potential energy function plays an important role in understanding the dynamics and function of complex molecules. We present a method to obtain path ensemble averages of a perturbed dynamics from a set of paths generated by a reference dynamics. It is based on the concept of path probability measure and the Girsanov theorem, a result from stochastic analysis to estimate a change of measure of a path ensemble. Since Markov state models (MSMs) of the molecular dynamics can be formulated as a combined phase-space and path ensemble average, the method can be extended to reweight MSMs by combining it with a reweighting of the Boltzmann distribution. We demonstrate how to efficiently implement the Girsanov reweighting in a molecular dynamics simulation program by calculating parts of the reweighting factor "on the fly" during the simulation, and we benchmark the method on test systems ranging from a two-dimensional diffusion process and an artificial many-body system to alanine dipeptide and valine dipeptide in implicit and explicit water. The method can be used to study the sensitivity of molecular dynamics on external perturbations as well as to reweight trajectories generated by enhanced sampling schemes to the original dynamics.

Neuroinvasive St. Louis Encephalitis Virus Infection in Solid Organ Transplant Recipients.

In summer 2015, three unrelated solid organ transplant recipients in Phoenix, Arizona, had meningoencephalitis suggestive of West Nile virus (WNV) infection. Testing was inconclusive but was later confirmed as St. Louis encephalitis (SLE). We retrospectively reviewed clinical manifestations, treatment, and outcomes of these transplant recipients. Common symptoms were fever, rigors, diarrhea, headache, and confusion. One patient died 3 days after hospitalization. Therapy for the other two patients was initiated with interferon α-2b (IFN) and intravenous IgG (IVIG; IFN plus IVIG in combination). Both patients tested positive for WNV by serologic assay, but SLE virus (SLEV) infection was later confirmed by plaque reduction neutralization test at a reference laboratory. Clinical improvement was observed within 72 h after initiation of IFN plus IVIG. SLEV has been an uncommon cause of neuroinvasive disease in the United States. Accurate, timely diagnosis is hindered because of clinical presentation similar to neuroinvasive WNV and SLE, serologic cross-reactivity, and lack of a commercially available serologic assay for SLEV. There is currently no approved therapy for flaviviral neuroinvasive disease. Anecdotal reports indicate varying success with IFN, IVIG, or IFN plus IVIG in WNV neuroinvasive disease. The same regimen might be of value for immunocompromised persons with neuroinvasive SLEV infection.

Mammary candidiasis: molecular-based detection of Candida species in human milk samples.

In this prospective and monocentric study, we investigated the performance of a commercialized real-time polymerase chain reaction (RT-PCR) test system for the specific detection of DNA from Candida albicans, C. dubliniensis, C. glabrata, C. krusei, C. lusitaniae, C. parapsilosis, and C. tropicalis in human milk samples of patients suspicious of mammary candidiasis. For this purpose, 43 breast-feeding women with characteristic symptoms of mammary candidiasis and 40 asymptomatic controls were enrolled. By culture, Candida spp. were detected in 8.8 % (4/46) and 9.3 % (4/43) of patient and control samples, respectively. Candida albicans (2/46), C. parapsilosis (1/46), and C. guilliermondii (1/46) were present in patient samples, and C. lusitaniae (3/43) and C. guilliermondii (1/43) were present in the controls. After RT-PCR was applied, Candida spp. were found to be present in 67.4 % (31/46) and 79.1 % (34/43) of patient and control samples investigated, respectively. PCR detection of C. albicans and C. parapsilosis revealed only a low sensitivity and specificity of 67.4 % and 41.9 %, respectively. Our data do not support the use of Candida RT-PCR for sensitive and specific diagnosis of mammary candidiasis.

Wnt9a deficiency discloses a repressive role of Tcf7l2 on endocrine differentiation in the embryonic pancreas.

Transcriptional and signaling networks establish complex cross-regulatory interactions that drive cellular differentiation during development. Using microarrays we identified the gene encoding the ligand Wnt9a as a candidate target of Neurogenin3, a basic helix-loop-helix transcription factor that functions as a master regulator of pancreatic endocrine differentiation. Here we show that Wnt9a is expressed in the embryonic pancreas and that its deficiency enhances activation of the endocrine transcriptional program and increases the number of endocrine cells at birth. We identify the gene encoding the endocrine transcription factor Nkx2-2 as one of the most upregulated genes in Wnt9a-ablated pancreases and associate its activation to reduced expression of the Wnt effector Tcf7l2. Accordingly, in vitro studies confirm that Tcf7l2 represses activation of Nkx2-2 by Neurogenin3 and inhibits Nkx2-2 expression in differentiated ß-cells. Further, we report that Tcf7l2 protein levels decline upon initiation of endocrine differentiation in vivo, disclosing the downregulation of this factor in the developing endocrine compartment. These findings highlight the notion that modulation of signalling cues by lineage-promoting factors is pivotal for controlling differentiation programs.

Long-term evaluation of impedance levels and clinical development in subthalamic deep brain stimulation for Parkinson's disease.

BACKGROUND: This study was conducted to better understand the development of clinical efficacy and impedance levels in the long-term course of deep brain stimulation (DBS) of the subthalamic nucleus (STN) in Parkinson's disease (PD). METHODS: In this retrospective study of twenty PD patients, the motor part of the Unified Parkinson's Disease Rating Scale was periodically assessed i) after withdrawal of medication and inactivated stimulation, ii) after withdrawal of medication with activated stimulation and iii) after challenge with l-Dopa during activated stimulation up to 13 years after surgery. RESULTS: STN-DBS with or without medication significantly improved motor function up to 13 years after surgery. The contribution of axial symptoms increased over time. While the stimulation parameters were kept constant, the therapeutic impedances progressively declined. CONCLUSION: STN-DBS in PD remains effective in the long-term course of the disease. Constant current stimulation might be preferable over voltage-controlled stimulation, as it would alleviate the impact of impedance changes on the volume of tissue activated.

Influence of pre-analytical procedures on genomic DNA integrity in blood samples: the SPIDIA experience.

BACKGROUND: DNA integrity is a critical part of the definition of genomic DNA (gDNA) quality and can influence downstream molecular applications. Pre-analytical variables as sample storage and DNA extraction methods can influence the quality and quantity of isolated DNA and affect molecular test performances. The aim of this paper is to investigate the role of blood sample storage and DNA extraction procedures on gDNA integrity and gDNA fragmentation impact on a molecular test. METHODS: 157 DNA samples deriving from the Pan European 1st SPIDIA DNA External Quality Assessment (EQA), aimed to investigate the influence of blood storage on gDNA quality and quantity, have been analyzed by Pulsed Field Gel Electrophoresis and ImageJ imaging software. 157 DNA samples derived from the Pan European 1st SPIDIA DNA External Quality Assessment (EQA), which aimed to investigate the influence of blood storage on gDNA quality and quantity, have been analyzed by Pulsed Field Gel Electrophoresis and ImageJ imaging software. RESULTS/CONCLUSIONS: Our results demonstrate that blood sample storage and DNA extraction procedures influence gDNA integrity and that the same blood sample which underwent a long range multiplex PCR based analytical test can provide different results if the adopted pre-analytical procedures are not standardized.

Adenomas and carcinomas missed in routine colonoscopy: a prospective study in resected colon segments.

Colonoscopy is the standard technique in the diagnosis and treatment of colorectal neoplasia, but small adenomas and even advanced lesions can be missed during the procedure. With large scale screening colonoscopy programs installed, information on quality of colonoscopy in primary care is essential, but scarcely available. Over a period of 45 months, we prospectively included all those patients in our study, who underwent major colonic surgery at our institution and who had undergone a colonoscopy within 42 days prior to the operation. 89 men and 100 women, median age 71 years, were included. The majority of these operations were performed for colorectal carcinoma (125), other malignant tumors (4), suspected malignancies (6) or large adenomas (14). The pathologist inspected the resected colonic segment, and we compared his findings with the colonoscopy report. Colonoscopies had been performed by 22 doctors in 13 institutions. Median length of the resected colonic segments was 20 cm (range 3 to 135 cm), total length was 41,21 metres. In 14 segments the pathologist identified 28 neoplastic lesions not described in the endoscopy report. Colonoscopy had missed 2 carcinomas, both in the right colon, and a 12 mm tubulo-villous adenoma with high-grade dysplasia. Another 25 tubular adenomas had been missed, 2 measuring 10 mm, 7 between 5 and 9 mm and 16 smaller than 5 mm. We conclude that primary care colonoscopy misses neoplastic lesions in a significant number of procedures. Most of the missed lesions in our high risk group of patients would have been of little clinical consequence. In a small, but clinically important number of cases, however, advanced adenomas and even colorectal carcinomas were missed by endoscopy.

NDRG1 prognosticates the natural course of disease in WHO grade II glioma.

There is a lack of relevant prognostic and predictive factors in neurooncology besides mutation of isocitrate dehydrogenase 1, codeletion of 1p/19q and promoter hypermethylation of O (6) -methylguanine-DNA-methyltransferase. More importantly, there is limited translation of these factors into clinical practice. The cancer genome atlas data and also clinical correlative analyses suggest a pivotal role for the epidermal growth factor receptor /protein kinase B/mammalian target of rapamycin (mTOR) pathway in both biology and the clinical course of gliomas. However, attempts to stratify gliomas by activating alterations in this pathway have failed thus far. The tumors of 40 patients with WHO grade II gliomas without immediate postoperative genotoxic treatment and known progression and survival status at a median follow-up of 12.2 years were analyzed for expression of the mTOR complex 2 downstream target N-myc downstream regulated gene (NDRG)1 using immunohistochemistry. Baseline characteristics for NDRG1 absent/low versus moderate/high patients were similar. Time to reintervention was significantly longer in the NDRG1 group (P = 0.026). NDRG1 may become a novel biomarker to guide the decision which WHO°II glioma patients may be followed without postsurgical intervention and which patients should receive genotoxic treatment early on. Validation of this hypothesis will be possible with the observational arm of the RTOG 9802 and the pretreatment step of the EORTC 22033/26032 trials.

SPIDIA-DNA: an External Quality Assessment for the pre-analytical phase of blood samples used for DNA-based analyses.

BACKGROUND: The EC-funded project SPIDIA is aimed to develop evidence-based quality guidelines for the pre-analytical phase of blood samples used for DNA molecular testing. To this purpose, a survey and a pan-European External Quality Assessment (EQA) were implemented. METHODS: SPIDIA facility sent to all the participants the same blood sample to be processed without time or temperature limitation. DNA quality parameters performed at SPIDIA facility included: UV spectrophotometric analysis of DNA purity and yield, PCR interferences study by Kineret software and DNA integrity analysis by pulsed field gel electrophoresis. RESULTS: 197 applications have been collected from 30 European countries. A high variability of DNA fragmentation was observed whereas purity, yield and PCR interferences had a narrow distribution within laboratories. A significant difference between the RNase P single copy gene quantity obtained in the DNA samples extracted with the precipitation-based method respect to those obtained with beads and column-based methods was observed. CONCLUSIONS: The results of this study will be the basis for implementing a second pan-European EQA and the results of both EQAs will be pooled and will provide the basis for the implementation of evidence-based guidelines for the pre-analytical phase of DNA analysis of blood samples.

Analysis of CIC-associated CpG island methylation in oligoastrocytoma.

AIMS: Combined deletion of the whole chromosomal arms 1p and 19q is a frequent event in oligodendroglial tumours. Recent identification of recurrent mutations in CIC on 19q and FUBP1 on 1p and their mutational patterns suggest a loss of function of the respective proteins. Surprisingly, oligoastrocytomas harbouring identical genetic characteristics regarding 1p/19q codeletion and frequent IDH1/2 mutations have been shown to carry CIC mutations in a significantly lower number of cases. The present study investigates whether epigenetic modification may result in silencing of CIC. METHODS: As IDH1/2 mutation-mediated DNA hypermethylation is a prominent feature of these tumours, we analysed a set of CIC wild-type oligoastrocytomas and other diffuse gliomas with regard to 1p/19q status for presence of CIC-associated CpG island methylation by methylation-specific PCR. RESULTS: Both methylation-specific PCR and subsequent bisulphite-sequencing of selected cases revealed an unmethylated status in all samples. CONCLUSION: Despite the hypermethylator phenotype in IDH1/2 mutant tumours and recent detection of gene silencing particularly on retained alleles in oligodendroglial tumours, hypermethylation of CIC-associated CpG islands does not provide an alternative mechanism of functional CIC protein abrogation.

Interlaboratory comparison of IDH mutation detection.

Isocitrate dehydrogenase (IDH) mutational testing is becoming increasingly important. For this, robust and reliable assays are needed. We tested the variation of results between six laboratories of testing for IDH mutations. Each laboratory received five unstained slides from 31 formalin-fixed paraffin-embedded (FFPE) glioma samples, and followed its own standard IDH diagnostic routine. All laboratories used immunohistochemistry (IHC) with an antibody against the most frequent IDH1 mutation (R132H) as a first step. Three laboratories then sequenced only IHC negative cases while the others sequenced all cases. Based on the overall analysis, 13 samples from 11 tumors had an R132H mutation and one tumor showed an R132G mutation. Results of IHC for IDH1 R132H mutations in all six laboratories were completely in agreement, and identified all R132H mutations. Upon sequencing the results of two laboratories deviated from those of the others. After a review of the entire diagnostic process, on repeat (blinded) testing one laboratory was completely in agreement with the overall result. A change in technique did only partially improve the results in the other laboratory. IHC for the IDH1 R132H mutation is very reliable and consistent across laboratories. IDH sequencing procedures yielded inconsistent results in 2 out of 6 laboratories. Quality assurance is pivotal before IDH testing is made part of clinical management of patients.