Development of Zalfermin, a Long-Acting Proteolytically Stabilized FGF21 Analog.

Here, we describe the development of the FGF21 analog zalfermin (NNC0194-0499, 15), intended for once-weekly sc dosing. Protein engineering was needed to address inherent druggability issues of the natural FGF21 hormone. Thus, deamidation of Asp121 was solved by mutation to glutamine, and oxidation of Met168 was solved by mutation to leucine. N-terminal region degradation by dipeptidyl peptidase IV was prevented by alanine residue elongation. To prevent inactivating metabolism by fibroblast activation protein and carboxypeptidase-like activity in the C-terminal region, and to achieve t1/2 extension (53 h in cynomolgus monkeys), we introduced a C18 fatty diacid at the penultimate position 180. The fatty diacid binds albumin in a reversible manner, such that the free fraction of zalfermin potently activates the FGF-receptor complex and retains receptor selectivity compared with FGF21, providing strong efficacy on body weight loss in diet-induced obese mice. Zalfermin is currently being clinically evaluated for the treatment of metabolic dysfunction-associated steatohepatitis.

A novel contaminant in museums? A cross-sectional study on xerophilic Aspergillus growth in climate-controlled repositories.

In the last decade, extensive fungal growth has developed in Danish museums parallel to climate change, challenging occupational health and heritage preservation. The growth was unexpected as the museums strived to control relative humidity below 60 %, and it should exceed 75 % to risk growth. A Danish case study found xerophilic Aspergillus species able to grow at low relative humidity in a museum repository. This cross-sectional study aimed to examine whether xerophilic growth from Aspergillus section Restricti has become a novel contaminant nationally distributed in Danish museum repositories striving to control relative humidity according to international environmental recommendations for heritage collections. The study examined The National Museum of Denmark and eight large State Recognized museums distributed throughout Denmark. It was based on 600 swab and tape-lift samples and 60 MAS100-Eco and filter air samples analyzed for fungi with cultivation and morphological identification, Big-Dye-Sanger sequencing, CaM-Nanopore and ITS-Illumina amplicon sequencing. The study showed growth from seven xerophilic Aspergillus species: A. conicus, A. domesticus, A. glabripes, A. halophilicus, A. magnivesiculatus, A. penicilloides, A. vitricola, of which three are new to Denmark, and 13 xerotolerant Aspergillus species. There was no growth from other fungal species. The multiple detection approach provided a broad characterization; however, there was variance in the detected species depending on the analysis approach. Cultivation and Big-Dye Sanger sequencing showed the highest Aspergillus diversity, detecting 17 species; CaM-Nanopore amplicon sequencing detected 12 species; and ITS-illumina amplicon sequencing detected two species but the highest overall diversity. Cultivation, followed by Big-Dye Sanger and CaM-amplicon sequencing, proved the highest compliance. The study concluded that xerophilic Aspergillus growth is nationally distributed and suggests species from Aspergillus section Restricti as a novel contaminant in climate-controlled museum repositories. To safeguard occupational health and heritage preservation research in sustainable solutions, avoiding xerophilic growth in museum collections is most important.

An efficient mRNA display protocol yields potent bicyclic peptide inhibitors for FGFR3c: outperforming linear and monocyclic formats in affinity and stability.

Macrocyclization has positioned itself as a powerful method for engineering potent peptide drug candidates. Introducing one or multiple cyclizations is a common strategy to improve properties such as affinity, bioavailability and proteolytic stability. Consequently, methodologies to create large libraries of polycyclic peptides by phage or mRNA display have emerged, allowing the rapid identification of binders to virtually any target. Yet, within those libraries, the performance of linear vs. mono- or bicyclic peptides has rarely been studied. Indeed, a key parameter to perform such a comparison is to use a display protocol and cyclization chemistry that enables the formation of all 3 formats in equal quality and diversity. Here, we developed a simple, efficient and fast mRNA display protocol which meets these criteria and can be used to generate highly diverse libraries of thioether cyclized polycyclic peptides. As a proof of concept, we selected peptides against fibroblast growth factor receptor 3c (FGFR3c) and compared the different formats regarding affinity, specificity, and human plasma stability. The peptides with the best KD's and stability were identified among bicyclic peptide hits, further strengthening the body of evidence pointing at the superiority of this class of molecules and providing functional and selective inhibitors of FGFR3c.

Subclinical epileptiform discharges in Alzheimer's disease are associated with increased hippocampal blood flow.

BACKGROUND: In epilepsy, the ictal phase leads to cerebral hyperperfusion while hypoperfusion is present in the interictal phases. Patients with Alzheimer's disease (AD) have an increased prevalence of epileptiform discharges and a study using intracranial electrodes have shown that these are very frequent in the hippocampus. However, it is not known whether there is an association between hippocampal hyperexcitability and regional cerebral blood flow (rCBF). The objective of the study was to investigate the association between rCBF in hippocampus and epileptiform discharges as measured with ear-EEG in patients with Alzheimer's disease. Our hypothesis was that increased spike frequency may be associated with increased rCBF in hippocampus. METHODS: A total of 24 patients with AD, and 15 HC were included in the analysis. Using linear regression, we investigated the association between rCBF as measured with arterial spin-labelling MRI (ASL-MRI) in the hippocampus and the number of spikes/sharp waves per 24 h as assessed by ear-EEG. RESULTS: No significant difference in hippocampal rCBF was found between AD and HC (p-value = 0.367). A significant linear association between spike frequency and normalized rCBF in the hippocampus was found for patients with AD (estimate: 0.109, t-value = 4.03, p-value < 0.001). Changes in areas that typically show group differences (temporal-parietal cortex) were found in patients with AD, compared to HC. CONCLUSIONS: Increased spike frequency was accompanied by a hemodynamic response of increased blood flow in the hippocampus in patients with AD. This phenomenon has also been shown in patients with epilepsy and supports the hypothesis of hyperexcitability in patients with AD. The lack of a significant difference in hippocampal rCBF may be due to an increased frequency of epileptiform discharges in patients with AD. TRIAL REGISTRATION: The study is registered at clinicaltrials.gov (NCT04436341).

Damp Buildings: Associated Fungi and How to Find Them.

The number of buildings experiencing humidity problems and fungal growth appears to be increasing as energy-saving measures and changes in construction practices and climate become more common. Determining the cause of the problem and documenting the type and extent of fungal growth are complex processes involving both building physics and indoor mycology. New detection and identification methods have been introduced, and new fungal species have been added to the list of building-related fungi. However, the lack of standardised procedures and general knowledge hampers the effort to resolve the problems and advocate for an effective renovation plan. This review provides a framework for building inspections on current sampling methods and detection techniques for building-related fungi. The review also contains tables with fungal species that have been identified on commonly used building materials in Europe and North America (e.g., gypsum wallboard, oriented strand board (OSB), concrete and mineral wool). The most reported building-associated fungi across all materials are Penicillium chrysogenum and Aspergillus versicolor. Chaetomium globosum is common on all organic materials, whereas Aspergillus niger is common on all inorganic materials.

Identification and engineering of potent cyclic peptides with selective or promiscuous binding through biochemical profiling and bioinformatic data analysis.

As our understanding of biological systems grows, so does the need to selectively target individual or multiple members of specific protein families in order to probe their function. Many targets of current biological and pharmaceutical interest are part of a large family of closely related proteins and achieving ligand selectivity often remains either an elusive or time-consuming endeavour. Cyclic peptides (CPs) occupy a key niche in ligand space, able to achieve high affinity and selectivity while retaining synthetic accessibility. De novo cyclic peptide ligands can be rapidly generated against a given target using mRNA display. In this study we harness mRNA display technology and the wealth of next generation sequencing (NGS) data generated to explore both experimental approaches and bioinformatic, statistical data analysis of peptide enrichment in cross-screen selections to rapidly generate high affinity CPs with differing intra-family protein selectivity profiles against fibroblast growth factor receptor (FGF-R) family proteins. Using these methods, CPs with distinct selectivity profiles can be generated which can serve as valuable tool compounds to decipher biological questions.

Copernicus for urban resilience in Europe.

The urban community faces a significant obstacle in effectively utilising Earth Observation (EO) intelligence, particularly the Copernicus EO program of the European Union, to address the multifaceted aspects of urban sustainability and bolster urban resilience in the face of climate change challenges. In this context, here we present the efforts of the CURE project, which received funding under the European Union's Horizon 2020 Research and Innovation Framework Programme, to leverage the Copernicus Core Services (CCS) in supporting urban resilience. CURE provides spatially disaggregated environmental intelligence at a local scale, demonstrating that CCS can facilitate urban planning and management strategies to improve the resilience of cities. With a strong emphasis on stakeholder engagement, CURE has identified eleven cross-cutting applications between CCS that correspond to the major dimensions of urban sustainability and align with user needs. These applications have been integrated into a cloud-based platform known as DIAS (Data and Information Access Services), which is capable of delivering reliable, usable and relevant intelligence to support the development of downstream services towards enhancing resilience planning of cities throughout Europe.

Subclinical Epileptiform Activity in Dementia with Lewy Bodies.

BACKGROUND: Patients with dementia with Lewy bodies (DLB) have a higher probability of seizures than in normal aging and in other types of neurodegenerative disorders. Depositions of α-synuclein, a pathological hallmark of DLB, can induce network excitability, which can escalate into seizure activity. Indicator of seizures are epileptiform discharges as observed using electroencephalography (EEG). However, no studies have so far investigated the occurrence of interictal epileptiform discharges (IED) in patients with DLB. OBJECTIVES: To investigate if IED as measured with ear-EEG occurs with a higher frequency in patients with DLB compared to healthy controls (HC). METHODS: In this longitudinal observational exploratory study, 10 patients with DLB and 15 HC were included in the analysis. Patients with DLB underwent up to three ear-EEG recordings, each lasting up to 2 days, over a period of 6 months. RESULTS: At baseline, IED were detected in 80% of patients with DLB and in 46.7% of HC. The spike frequency (spikes or sharp waves/24 hours) was significantly higher in patients with DLB as compared to HC with a risk ratio of 2.52 (CI, 1.42-4.61; P-value = 0.001). Most IED occurred at night. CONCLUSIONS: Long-term outpatient ear-EEG monitoring detects IED in most patients with DLB with an increased spike frequency compared to HC. This study extends the spectrum of neurodegenerative disorders in which epileptiform discharges occurs at an elevated frequency. It is possible that epileptiform discharges are, therefore, a consequence of neurodegeneration. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Renoprotective effects of genetically proxied fibroblast growth factor 21: Mendelian randomization, proteome-wide and metabolome-wide association study.

BACKGROUND: Fibroblast growth factor 21 (FGF21) has demonstrated efficacy for reducing liver fat and reversing non-alcoholic steatohepatitis in phase 2 clinical trials. It is also postulated to have anti-fibrotic effects and therefore may be amenable to repurposing for the prevention and treatment of chronic kidney disease (CKD). METHODS: We leverage a missense genetic variant, rs739320 in the FGF21 gene, that associates with magnetic resonance imaging-derived liver fat as a clinically validated and biologically plausible instrumental variable for studying the effects of FGF21 analogs. Performing Mendelian randomization, we ascertain associations between instrumented FGF21 and kidney phenotypes, cardiometabolic disease risk factors, as well as the circulating proteome (Somalogic, 4907 aptamers) and metabolome (Nightingale platform, 249 metabolites). RESULTS: We report consistent renoprotective associations of genetically proxied FGF21 effect, including higher glomerular filtration rates (p = 1.9 × 10-4), higher urinary sodium excretion (p = 5.1 × 10-11), and lower urine albumin-creatinine ratio (p = 3.6 × 10-5). These favorable effects translated to lower CKD risk (odds ratio per rs739320 C-allele, 0.96; 95%CI, 0.94-0.98; p = 3.2 × 10-4). Genetically proxied FGF21 effect was also associated with lower fasting insulin, waist-to-hip ratio, blood pressure (systolic and diastolic BP, p < 1.0 × 10-07) and blood lipid (low-density lipoprotein cholesterol, triglycerides and apolipoprotein B, p < 6.5 × 10-24) profiles. The latter associations are replicated in our metabolome-wide association study. Proteomic perturbations associated with genetically predicted FGF21 effect were consistent with fibrosis reduction. CONCLUSION: This study highlights the pleiotropic effects of genetically proxied FGF21 and supports a re-purposing opportunity for the treatment and prevention of kidney disease specifically. Further work is required to triangulate these findings, towards possible clinical development of FGF21 towards the treatment and prevention of kidney disease.

Hepatocyte mARC1 promotes fatty liver disease.

Background & Aims: Non-alcoholic fatty liver disease (NAFLD) has a prevalence of ∼25% worldwide, with significant public health consequences yet few effective treatments. Human genetics can help elucidate novel biology and identify targets for new therapeutics. Genetic variants in mitochondrial amidoxime-reducing component 1 (MTARC1) have been associated with NAFLD and liver-related mortality; however, its pathophysiological role and the cell type(s) mediating these effects remain unclear. We aimed to investigate how MTARC1 exerts its effects on NAFLD by integrating human genetics with in vitro and in vivo studies of mARC1 knockdown. Methods: Analyses including multi-trait colocalisation and Mendelian randomisation were used to assess the genetic associations of MTARC1. In addition, we established an in vitro long-term primary human hepatocyte model with metabolic readouts and used the Gubra Amylin NASH (GAN)-diet non-alcoholic steatohepatitis mouse model treated with hepatocyte-specific N-acetylgalactosamine (GalNAc)-siRNA to understand the in vivo impacts of MTARC1. Results: We showed that genetic variants within the MTARC1 locus are associated with liver enzymes, liver fat, plasma lipids, and body composition, and these associations are attributable to the same causal variant (p.A165T, rs2642438 G>A), suggesting a shared mechanism. We demonstrated that increased MTARC1 mRNA had an adverse effect on these traits using Mendelian randomisation, implying therapeutic inhibition of mARC1 could be beneficial. In vitro mARC1 knockdown decreased lipid accumulation and increased triglyceride secretion, and in vivo GalNAc-siRNA-mediated knockdown of mARC1 lowered hepatic but increased plasma triglycerides. We found alterations in pathways regulating lipid metabolism and decreased secretion of 3-hydroxybutyrate upon mARC1 knockdown in vitro and in vivo. Conclusions: Collectively, our findings from human genetics, and in vitro and in vivo hepatocyte-specific mARC1 knockdown support the potential efficacy of hepatocyte-specific targeting of mARC1 for treatment of NAFLD. Impact and implications: We report that genetically predicted increases in MTARC1 mRNA associate with poor liver health. Furthermore, knockdown of mARC1 reduces hepatic steatosis in primary human hepatocytes and a murine NASH model. Together, these findings further underscore the therapeutic potential of targeting hepatocyte MTARC1 for NAFLD.

Detection of subclinical epileptiform discharges in Alzheimer's disease using long-term outpatient EEG monitoring.

BACKGROUND: In patients with Alzheimer's disease (AD) without clinical seizures, up to half have epileptiform discharges on long-term in-patient electroencephalography (EEG) recordings. Long-term in-patient monitoring is obtrusive, and expensive as compared to outpatient monitoring. No studies have so far investigated if long-term outpatient EEG monitoring is able to identify epileptiform discharges in AD. Our aim is to investigate if epileptiform discharges as measured with ear-EEG are more common in patients with AD compared to healthy elderly controls (HC). METHODS: In this longitudinal observational study, 24 patients with mild to moderate AD and 15 age-matched HC were included in the analysis. Patients with AD underwent up to three ear-EEG recordings, each lasting up to two days, within 6 months. RESULTS: The first recording was defined as the baseline recording. At baseline, epileptiform discharges were detected in 75.0% of patients with AD and in 46.7% of HC (p-value = 0.073). The spike frequency (spikes or sharp waves/24 h) was significantly higher in patients with AD as compared to HC with a risk ratio of 2.90 (CI: 1.77-5.01, p < 0.001). Most patients with AD (91.7%) showed epileptiform discharges when combining all ear-EEG recordings. CONCLUSIONS: Long-term ear-EEG monitoring detects epileptiform discharges in most patients with AD with a three-fold increased spike frequency compared to HC, which most likely originates from the temporal lobes. Since most patients showed epileptiform discharges with multiple recordings, elevated spike frequency should be considered a marker of hyperexcitability in AD.

Aerobic exercise does not affect serum neurofilament light in patients with mild Alzheimer's disease.

Introduction: Aerobic exercise has been shown to modify Alzheimer pathology in animal models, and in patients with multiple sclerosis to reduce neurofilament light (NfL), a biomarker of neurodegeneration. Objective: To investigate whether a 16-week aerobic exercise program was able to reduce serum NfL in patients with mild Alzheimer's disease (AD). Methods: This is a secondary analysis of data from the multi-center Preserving Cognition, Quality of Life, Physical Health, and Functional Ability in Alzheimer's disease: The Effect of Physical Exercise (ADEX) study. Participants were randomized to 16 weeks of moderate intensity aerobic exercise or usual care. Clinical assessment and measurement of serum NfL was done at baseline and after the intervention. Results: A total of 136 participants were included in the analysis. Groups were comparable at baseline except for APOEε4 carriership which was higher in the usual care group (75.3 versus 60.2%; p = 0.04). There was no effect of the intervention on serum NfL [intervention: baseline NfL (pg/mL) 25.76, change from baseline 0.87; usual care: baseline 27.09, change from baseline -1.16, p = 0.09]. Conclusion: The findings do not support an effect of the exercise intervention on a single measure of neurodegeneration in AD. Further studies are needed using other types and durations of exercise and other measures of neurodegeneration. Clinical trial registration: clinicaltrials.gov, identifier NCT01681602.

Quantitative flow ratio versus fractional flow reserve for guiding percutaneous coronary intervention: design and rationale of the randomised FAVOR III Europe Japan trial.

Quantitative flow ratio (QFR) is a computation of fractional flow reserve (FFR) based on invasive coronary angiographic images. Calculating QFR is less invasive than measuring FFR and may be associated with lower costs. Current evidence supports the call for an adequately powered randomised comparison of QFR and FFR for the evaluation of intermediate coronary stenosis. The aim of the FAVOR III Europe Japan trial is to investigate if a QFR-based diagnostic strategy yields a non-inferior 12-month clinical outcome compared with a standard FFR-guided strategy in the evaluation of patients with intermediary coronary stenosis. FAVOR III Europe Japan is an investigator-initiated, randomised, clinical outcome, non-inferiority trial scheduled to randomise 2,000 patients with either 1) stable angina pectoris and intermediate coronary stenosis, or 2) indications for functional assessment of at least 1 non-culprit lesion after acute myocardial infarction. Up to 40 international centres will randomise patients to either a QFR-based or a standard FFR-based diagnostic strategy. The primary endpoint of major adverse cardiovascular events is a composite of all-cause mortality, any myocardial infarction, and any unplanned coronary revascularisation at 12 months. QFR could emerge as an adenosine- and wire-free alternative to FFR, making the functional evaluation of intermediary coronary stenosis less invasive and more cost-effective.

Predictive value of post-percutaneous coronary intervention fractional flow reserve: a systematic review and meta-analysis.

AIMS: We aimed to investigate the relationship between post-percutaneous coronary intervention (PCI) fractional flow reserve (FFR) and clinical outcome using a systematic review with a study-level meta-analysis. METHODS AND RESULTS: MEDLINE, Embase, and CENTRAL were systematically searched for articles with clinical follow-up reporting mean or median final post-PCI FFR. The main outcome was a composite of major adverse cardiac events (MACE) including all-cause death, myocardial infarction (MI), and target vessel revascularization (TVR). Meta-regression analyses were performed on mean post-PCI FFR values. A total of 62 studies with 12 340 patients and 12 923 stented vessels were included, with follow-ups ranging from 1 to 89 months. Post-PCI FFR was not continuously associated with the rate of 1-year MACE or 1-year TVR using meta-regression models accounting for heterogeneous follow-up lengths. For studies comparing high vs. low post-PCI FFR, low post-PCI FFR was associated with high risk ratio for MACE {1.97 [95% confidence interval (CI):1.45-2.67]}, all-cause death [1.59 (95% CI: 1.08-2.34)], MI [3.18 (95% CI: 1.84-5.50)], TVR [2.08 (95% CI: 1.63-2.65)] and angina status [2.50 (95% CI: 1.53-4.06)] using different optimal cut-off values spanning from 0.80 to 0.95. CONCLUSION: We found no clear continuous association between post-PCI FFR and clinical outcomes in this systematic study-level meta-analysis. In a subset of studies investigating binary classification, high post-PCI FFR was associated with a better clinical outcome than low post-PCI FFR.We investigated the relationship between post-percutaneous coronary intervention (PCI) fractional flow reserve (FFR) and rate of major adverse cardiac events (MACE), including all-cause death, myocardial infarction (MI), and target vessel revascularization (TVR), using a systematic review and study-level meta-analysis, pooling 12 340 patients from 62 studies. Mean post-PCI FFR was not continuously associated with a 1-year MACE rate accounting for heterogenous follow-up lengths. Still, the risk ratio favoured high post-PCI FFR for reduced MACE, all-cause death, MI, TVR, and better angina status using different cut-offs.

Long-Term EEG Monitoring in Patients with Alzheimer's Disease Using Ear-EEG: A Feasibility Study.

BACKGROUND: Previous studies have reported that epileptiform activity may be detectible in nearly half of patients with Alzheimer's disease (AD) on long-term electroencephalographic (EEG) recordings. However, such recordings can be uncomfortable, expensive, and difficult. Ear-EEG has shown promising results for long-term EEG monitoring, but it has not been used in patients with AD. OBJECTIVE: To investigate if ear-EEG is a feasible method for long-term EEG monitoring in patients with AD. METHODS: In this longitudinal, single-group feasibility study, ten patients with mild to moderate AD were recruited. A total of three ear-EEG recordings of up to 48 hours three months apart for six months were planned. RESULTS: All patients managed to wear the ear-EEG for at least 24 hours and at least one full night. A total of 19 ear-EEG recordings were performed (self-reported recording, mean: 37.15 hours (SD: 8.96 hours)). After automatic pre-processing, a mean of 27.37 hours (SD: 7.19 hours) of data with acceptable quality in at least one electrode in each ear was found. Seven out of ten participants experienced mild adverse events. Six of the patients did not complete the study with three patients not wanting to wear the ear-EEG anymore due to adverse events. CONCLUSION: It is feasible and safe to use ear-EEG for long-term EEG monitoring in patients with AD. Minor adjustments to the equipment may improve the comfort for the participants.

Challenges faced by people with a stoma: peristomal body profile risk factors and leakage.

AIM: The Ostomy Life Study 2019 aimed to obtain a better understanding of the challenges faced by people with stoma. METHODS: Online survey with participants from 17 countries. FINDINGS: Of the 54 614 individuals invited to take part, 5187 responded; 62% of the respondents avoided physical and social activities because of their stoma and 37% had never consulted their stoma care nurse to have the fit of their stoma product checked. In a subgroup receiving questions on leakage (n=4209), output under the baseplate and leakage onto clothes were experienced within the previous month by 76% and 26% of respondents, respectively. Higher chance of leakage was associated with an irregular stoma shape and peristomal body profile; a stoma level at or below the skin surface; and the presence of creases, folds and other changes in the peristomal area. CONCLUSION: Leakage and access to a stoma care nurse to provide the necessary care and guidance remain important concerns for individuals with a stoma.

The Ostomy Skin Tool 2.0: a new instrument for assessing peristomal skin changes.

BACKGROUND: Peristomal skin complications (PSCs) are frequently reported postoperative complications. PSCs can present visibly or as symptoms such as pain, itching or burning sensations. AIM: To develop a new tool that can capture a range of sensation symptoms together with visible complications and an objective assessment of discolouration in the peristomal area. METHOD: Consensus from qualitative interviews with health professionals and people with an ostomy, and input from expert panels, formed the basis of a patient-reported outcome (PRO) questionnaire. A decision tree model was used to define a combined score including PRO and objectively assessed discolouration area. FINDINGS: Six elements were included in the PRO questionnaire and four health states representing different severity levels of the peristomal skin were defined. CONCLUSION: The Ostomy Skin Tool 2.0 is a sensitive tool that can be used to follow changes in the peristomal skin on a regular basis and thereby help prevent severe PSCs.

Reproducibility of quantitative flow ratio: the QREP study.

BACKGROUND: Quantitative flow ratio (QFR) is a tool for physiological lesion assessment based on invasive coronary angiography. AIMS: We aimed to assess the reproducibility of QFR computed from the same angiograms as assessed by multiple observers from different, international sites. METHODS: We included 50 patients previously enrolled in dedicated QFR studies. QFR was computed twice, one month apart by five blinded observers. The main analysis was the coefficient of variation (CV) as a measure of intra- and inter-observer reproducibility. Key secondary analysis was the identification of clinical and procedural characteristics predicting reproducibility. RESULTS: The intra-observer CV ranged from 2.3% (1.5-2.8) to 10.2% (6.6-12.0) among the observers. The inter-observer CV was 9.4% (8.0-10.5). The QFR observer, low angiographic quality, and low fractional flow reserve (FFR) were independent predictors of a large absolute difference between repeated QFR measurements defined as a difference larger than the median difference (>0.03). CONCLUSIONS: The inter- and intra-observer reproducibility for QFR computed from the same angiograms ranged from high to poor among multiple observers from different sites with an average agreement of 0.01±0.08 for repeated measurements. The reproducibility was dependent on the observer, angiographic quality and the coronary artery stenosis severity as assessed by FFR.

Alternaria in malting barley: Characterization and distribution in relation with climatic conditions and barley cultivars.

Alternaria is one of the main fungal genera affecting the quality of barley grains. In this study, a polyphasic approach was carried out to characterise the Alternaria population infecting different cultivars of barley grains from the major producing regions of Argentina in the 2014 and 2015 seasons. Its relationship with Fusarium and correlations between predominant species, barley cultivars, and climatic conditions in the growing regions were evaluated. Alternaria incidence exceeded that of Fusarium in all the barley samples and was higher in the drier season (21% in 2014 and 42% in 2015 vs. 6% and 4%, respectively). The main Alternaria species-groups identified were present in both growing seasons in similar frequencies (A. tenuissima sp.-grp., 83.4% in 2014 and 81.7% in 2015; A. infectoria sp.-grp., 11.7% in 2014 and 11.3% in 2015). The dominant Alternaria species-group isolated and identified based on morphological characteristics, DNA sequencing, and metabolite profile was A. tenuissima (72.9%), followed by A. infectoria (14.6%). An association between their frequency and field temperature was observed; A. tenuissima sp.-grp. was more frequent in northern localities, where higher temperatures were registered, while the opposite was observed for A. infectoria sp.-grp. A smaller percentage of A. arborescens sp.-grp. (5%), A. alternata sp.-grp. (3.9%) and A. vaccinii (1.4%) were also identified. Both secondary metabolite profiles and phylogenetic analysis were useful to distinguish isolates from Alternaria section Alternaria and section Infectoriae. Regarding metabolite profiles, alternariol was the most frequent compound produced by isolates of the section Alternaria. Infectopyrones and novae-zelandins were produced by most of the isolates from section Infectoriae. The barley cultivars analysed in this study did not show a particular susceptibility regarding the Alternaria population composition, except for Andreia, which presented the highest frequency of contamination with A. tenuissima sp.-grp. The rest of the cultivars, when grown in different regions, showed different proportion of the Alternaria sp.-grps., suggesting that other factors were determinant in their distribution. The results obtained in the present study will be a valuable tool for health authorities to assess the need for regulations on Alternaria mycotoxins, given the high incidence of Alternaria spp. in barley and the diversity of metabolites that might contaminate the grains.

A pyrexic effect of FGF21 independent of energy expenditure and UCP1.

OBJECTIVE: Administration of FGF21 to mice reduces body weight and increases body temperature. The increase in body temperature is generally interpreted as hyperthermia, i.e. a condition secondary to the increase in energy expenditure (heat production). Here, we examine an alternative hypothesis: that FGF21 has a direct pyrexic effect, i.e. FGF21 increases body temperature independently of any effect on energy expenditure. METHODS: We studied the effects of FGF21 treatment on body temperature and energy expenditure in high-fat-diet-fed and chow-fed mice exposed acutely to various ambient temperatures, in high-fat diet-fed mice housed at 30 °C (i.e. at thermoneutrality), and in mice lacking uncoupling protein 1 (UCP1). RESULTS: In every model studied, FGF21 increased body temperature, but energy expenditure was increased only in some models. The effect of FGF21 on body temperature was more (not less, as expected in hyperthermia) pronounced at lower ambient temperatures. Effects on body temperature and energy expenditure were temporally distinct (daytime versus nighttime). FGF21 enhanced UCP1 protein content in brown adipose tissue (BAT); there was no measurable UCP1 protein in inguinal brite/beige adipose tissue. FGF21 increased energy expenditure through adrenergic stimulation of BAT. In mice lacking UCP1, FGF21 did not increase energy expenditure but increased body temperature by reducing heat loss, e.g. a reduced tail surface temperature. CONCLUSION: The effect of FGF21 on body temperature is independent of UCP1 and can be achieved in the absence of any change in energy expenditure. Since elevated body temperature is a primary effect of FGF21 and can be achieved without increasing energy expenditure, only limited body weight-lowering effects of FGF21 may be expected.