Discovery and Characterization of Novel Vascular and Hematopoietic Genes Downstream of Etsrp in Zebrafish

The transcription factor Etsrp is required for vasculogenesis and primitive myelopoiesis in zebrafish. When ectopically expressed, etsrp is sufficient to induce the expression of many vascular and myeloid genes in zebrafish. The mammalian homolog of etsrp, ER71/Etv2, is also essential for vascular and hematopoietic development. To identify genes downstream of etsrp, gain-of-function experiments were performed for etsrp in zebrafish embryos followed by transcription profile analysis by microarray. Subsequent in vivo expression studies resulted in the identification of fourteen genes with blood and/or vascular expression, six of these being completely novel. Regulation of these genes by etsrp was confirmed by ectopic induction in etsrp overexpressing embryos and decreased expression in etsrp deficient embryos. Additional functional analysis of two newly discovered genes, hapln1b and sh3gl3, demonstrates their importance in embryonic vascular development. The results described here identify a group of genes downstream of etsrp likely to be critical for vascular and/or myeloid development

Total Lipids and Fatty Acids in Major New Zealand Grape Varieties during Ripening, Prolonged Pomace Contacts and Ethanolic Extractions Mimicking Fermentation

Despite the important roles of lipids in winemaking, changes in lipids during grape ripening are largely unknown for New Zealand (NZ) varieties. Therefore, we aimed to determine the fatty acid profiles and total lipid content in two of NZ’s major grape varieties. Using gas chromatography–mass spectrometry, absolute quantification of 45 fatty acids was determined in Sauvignon blanc (SB) and Pinot noir (PN) grapes harvested at two different stages of ripeness. Lipid concentrations were as high as 0.4 g/g in seeds of both varieties, while pulp contained the least amount. Many unsaturated fatty acids were present, particularly in grape seeds, while skin contained relatively higher amounts of saturated fatty acids that increased throughout ripening. For both varieties, a significant increase in lipid concentration was observed in grapes harvested at the later stage of ripeness, indicating an association between lipids and grape maturity, and providing a novel insight about the use of total lipids as another parameter of grape ripeness. A variety-specific trend in the development and extraction of grape lipids was found from the analysis of the must and ethanolic extracts. Lipid extraction increased linearly with the ethanol concentration and with the extended pomace contact time. More lipids were extracted from the SB pomace to the must than PN within 144 h, suggesting a must matrix effect on lipid extraction. The knowledge generated here is relevant to both industry and academia and can be used to develop lipid diversification strategies to produce different wine styles

Fully Automated Trimethylsilyl (TMS) Derivatisation Protocol for Metabolite Profiling by GC-MS

Gas Chromatography-Mass Spectrometry (GC-MS) has long been used for metabolite profiling of a wide range of biological samples. Many derivatisation protocols are already available and among these, trimethylsilyl (TMS) derivatisation is one of the most widely used in metabolomics. However, most TMS methods rely on off-line derivatisation prior to GC-MS analysis. In the case of manual off-line TMS derivatisation, the derivative created is unstable, so reduction in recoveries occurs over time. Thus, derivatisation is carried out in small batches. Here, we present a fully automated TMS derivatisation protocol using robotic autosamplers and we also evaluate a commercial software, Maestro available from Gerstel GmbH. Because of automation, there was no waiting time of derivatised samples on the autosamplers, thus reducing degradation of unstable metabolites. Moreover, this method allowed us to overlap samples and improved throughputs. We compared data obtained from both manual and automated TMS methods performed on three different matrices, including standard mix, wine, and plasma samples. The automated TMS method showed better reproducibility and higher peak intensity for most of the identified metabolites than the manual derivatisation method. We also validated the automated method using 114 quality control plasma samples. Additionally, we showed that this online method was highly reproducible for most of the metabolites detected and identified (RSD < 20) and specifically achieved excellent results for sugars, sugar alcohols, and some organic acids. To the very best of our knowledge, this is the first time that the automated TMS method has been applied to analyse a large number of complex plasma samples. Furthermore, we found that this method was highly applicable for routine metabolite profiling (both targeted and untargeted) in any metabolomics laboratory

Factors Influencing Postgraduate Career Decisions of Ophthalmology Residents

AbstractObjective This study aims to identify factors that influence ophthalmology residents' decision to pursue fellowship training or to practice comprehensive ophthalmology after residency.Design This is a cross-sectional study.Methods An anonymous survey was sent to ophthalmology residents in the United States from the graduating class of 2018. The main outcome measure was the decision to seek fellowship training or to practice comprehensive ophthalmology. Information on demographics, residency program characteristics, and factors influencing career choices were collected.Results The overall response rate was 24.0% (112/467). Among the 112 respondents, 88 (78.6%) matched into subspecialty training. Compared with residents entering comprehensive ophthalmology, there was a greater proportion of Asians (37.5 vs. 8.3%) and a smaller proportion of non-Hispanic whites (47.7 vs. 70.8%,p = 0.029) among residents pursuing fellowship training. Residents pursuing fellowships had fewer children (0.4 vs. 0.8,p = 0.049), had less debt (median debt ≤ $100,000 vs. >$200,000,p = 0.008), had more first-author publications (4.3 vs. 1.2,p< 0.001), decided on their postgraduate career path earlier (median time postgraduate second year [PGY2] versus postgraduate third year [PGY3],p = 0.016), and were more likely to plan to practice in an academic setting (51.1 vs. 29.2%,p< 0.001) and in an urban location (44.3 vs. 12.5%,p = 0.004). In a multiple logistic regression model, factors predictive of fellowship training included a desire to acquire special skills (odds ratio [OR] =4.39; 95% confidence intervals [CI]: 1.70–16.37) and work with new technology (OR = 2.92; 95% CI: 1.16–10.02). Factors that predicted a career in comprehensive ophthalmology were higher levels of educational debt (OR = 0.35; 95% CI: 0.12–0.77), a later timing of postgraduate decision (OR = 0.17; 95% CI: 0.02–0.65), and lifestyle considerations (OR = 0.13; 95% CI: 0.03–0.36). Gender, prestige, or perceived favorable job market was not significant factors.Conclusion The majority of U.S. ophthalmology residents matched into fellowships. A desire to gain special skills and a desire to work with new technology were major factors influencing residents to seek fellowship training. Lifestyle considerations and educational debt were more important in the decision to choose a comprehensive ophthalmology career. Future research that includes more trainees and programs may minimize the selection bias issues present in this study

Individuals with and without normal tension glaucoma exhibit comparable performance on tests of cognitive function

AimTo evaluate aspects of cognition impacted by individuals with and without normal tension glaucoma.MethodsFifty normal tension glaucoma (NTG) and 50 control patients ≥50y of age were recruited from the UCSF Department of Ophthalmology. Demographic data and glaucoma parameters were extracted from electronic medical records for both groups. Tests of executive function [Executive Abilities: Measures and Instruments for Neurobehavioral Evaluation and Research (EXAMINER)] and learning and memory [California Verbal Learning Test-Second Edition (CVLT-II)] were administered to both NTG and controls. Race, handedness, best-corrected visual acuity, maximum intraocular pressure, optic nerve cup-to-disc ratio, visual field and optic nerve optical coherence tomography parameters, and a measure of general health (Charlson Comorbidity Index) were compared between NTG and controls as well as within NTG subgroups. Multivariate linear regression was used to compare group performances on the EXAMINER battery and CVLT-II while controlling for age, sex, and years of education.ResultsNTG and controls were comparable with respect to age, sex, race, education, handedness, and the Charlson Comorbidity Index (P&gt;0.05 for all). Performance on the EXAMINER composite score and the CVLT-II did not differ between NTG and controls (P&gt;0.05 for both).ConclusionThis is the first prospective study in which the cognitive function of subject with NTG were evaluated using a comprehensive, computerized neurocognitive battery. Subjects with NTG do not perform worse than unaffected controls on tests of executive function, learning, and memory. Results do not support the hypothesis that individuals with NTG are at higher risk for cognitive dysfunction and/or dementia

Metabolomics and a Breath Sensor Identify Acetone as a Biomarker for Heart Failure

Background: Multi-omics delivers more biological insight than targeted investigations. We applied multi-omics to patients with heart failure with reduced ejection fraction (HFrEF). Methods: 46 patients with HFrEF and 20 controls underwent metabolomic profiling, including liquid/gas chromatography mass spectrometry (LC-MS/GC-MS) and solid-phase microextraction (SPME) volatilomics in plasma and urine. HFrEF was defined using left ventricular global longitudinal strain, ejection fraction and NTproBNP. A consumer breath acetone (BrACE) sensor validated results in n = 73. Results: 28 metabolites were identified by GCMS, 35 by LCMS and 4 volatiles by SPME in plasma and urine. Alanine, aspartate and glutamate, citric acid cycle, arginine biosynthesis, glyoxylate and dicarboxylate metabolism were altered in HFrEF. Plasma acetone correlated with NT-proBNP (r = 0.59, 95% CI 0.4 to 0.7), 2-oxovaleric and cis-aconitic acid, involved with ketone metabolism and mitochondrial energetics. BrACE > 1.5 ppm discriminated HF from other cardiac pathology (AUC 0.8, 95% CI 0.61 to 0.92, p < 0.0001). Conclusion: Breath acetone discriminated HFrEF from other cardiac pathology using a consumer sensor, but was not cardiac specific