Delta-Notch signaling and lateral inhibition in zebrafish spinal cord development

BACKGROUND: Vertebrate neural development requires precise coordination of cell proliferation and cell specification to guide orderly transition of mitotically active precursor cells into different types of post-mitotic neurons and glia. Lateral inhibition, mediated by the Delta-Notch signaling pathway, may provide a mechanism to regulate proliferation and specification in the vertebrate nervous system. We examined delta and notch gene expression in zebrafish embryos and tested the role of lateral inhibition in spinal cord patterning by ablating cells and genetically disrupting Delta-Notch signaling. RESULTS: Zebrafish embryos express multiple delta and notch genes throughout the developing nervous system. All or most proliferative precursors appeared to express notch genes whereas subsets of precursors and post-mitotic neurons expressed delta genes. When we ablated identified primary motor neurons soon after they were born, they were replaced, indicating that specified neurons laterally inhibit neighboring precursors. Mutation of a delta gene caused precursor cells of the trunk neural tube to cease dividing prematurely and develop as neurons. Additionally, mutant embryos had excess early specified neurons, with fates appropriate for their normal positions within the neural tube, and a concomitant deficit of late specified cells. CONCLUSIONS: Our results are consistent with the idea that zebrafish Delta proteins, expressed by newly specified neurons, promote Notch activity in neighboring precursors. This signaling is required to maintain a proliferative precursor population and generate late-born neurons and glia. Thus, Delta-Notch signaling may diversify vertebrate neural cell fates by coordinating cell cycle control and cell specification

Absorption and metabolism of modified mycotoxins of alternariol, alternariol monomethyl ether, and zearalenone in Caco-2 cells

Background and objectives Various cereals, fruits, and vegetables are commonly contaminated with mycotoxins such as zearalenone (ZEN), alternariol (AOH), and alternariol monomethyl ether (AME). More recently, their glucosidic metabolites formed in the plant have gained increasing attention. Experimental data on the contribution of modified mycotoxins to total toxicity are either controversy or lacking. Thus, the aim of this study was to investigate the absorption and metabolism of ZEN‐, AOH‐, and AME‐glucosides using the Caco‐2 cell system. Findings No quantifiable amounts of ZEN‐14‐glucoside, ZEN‐16‐glucoside, free ZEN, and ZEN metabolites were found in Caco‐2 cells and in the basolateral compartment. In contrast, glucosides of AOH and AME were absorbed and released their parental toxins, which were further metabolized to form glucuronides and sulfates to a variable extent. Metabolites were found on the basolateral site, too. There is also evidence that AOH‐9‐diglucoside is hydrolyzed to AOH‐9‐glucoside. Conclusion Our results demonstrate that modified ZEN is less absorbed whereas modified Alternaria toxins are taken up to a higher extent by Caco‐2 cells, followed by deglucosylation, metabolization, and transport to the basolateral site, suggesting a potential contribution to overall toxicity of these modified mycotoxins. Significance and novelty For the first time, absorption studies using modified Alternaria toxins in the Caco‐2 cell system were carried out

Highly sensitive real-time PCR for specific detection and quantification of Coxiella burnetii

BACKGROUND: Coxiella burnetii, the bacterium causing Q fever, is an obligate intracellular biosafety level 3 agent. Detection and quantification of these bacteria with conventional methods is time consuming and dangerous. During the last years, several PCR based diagnostic assays were developed to detect C. burnetii DNA in cell cultures and clinical samples. We developed and evaluated TaqMan-based real-time PCR assays that targeted the singular icd (isocitrate dehydrogenase) gene and the transposase of the IS1111a element present in multiple copies in the C. burnetii genome. RESULTS: To evaluate the precision of the icd and IS1111 real-time PCR assays, we performed different PCR runs with independent DNA dilutions of the C. burnetii Nine Mile RSA493 strain. The results showed very low variability, indicating efficient reproducibility of both assays. Using probit analysis, we determined that the minimal number of genome equivalents per reaction that could be detected with a 95% probability was 10 for the icd marker and 6.5 for the IS marker. Plasmid standards with cloned icd and IS1111 fragments were used to establish standard curves which were linear over a range from 10 to 10(7 )starting plasmid copy numbers. We were able to quantify cell numbers of a diluted, heat-inactivated Coxiella isolate with a detection limit of 17 C. burnetii particles per reaction. Real-time PCR targeting both markers was performed with DNA of 75 different C. burnetii isolates originating from all over the world. Using this approach, the number of IS1111 elements in the genome of the Nine Mile strain was determined to be 23, close to 20, the number revealed by genome sequencing. In other isolates, the number of IS1111 elements varied widely (between seven and 110) and seemed to be very high in some isolates. CONCLUSION: We validated TaqMan-based real-time PCR assays targeting the icd and IS1111 markers of C. burnetii. The assays were shown to be specific, highly sensitive and efficiently reproducible. Cell numbers in dilutions of a C. burnetii isolate were reliably quantified. PCR quantification suggested a high variability of the number of IS1111 elements in different C. burnetii isolates, which may be useful for further phylogenetic studies

Reduction of aversive learning rates in Pavlovian conditioning by angiotensin II antagonist losartan: a randomized controlled trial

Background: Angiotensin receptor blockade has been linked to aspects of aversive learning and memory formation and to the prevention of posttraumatic stress disorder symptom development. Methods: We investigated the influence of the angiotensin receptor blocker losartan on aversive Pavlovian conditioning using a probabilistic learning paradigm. In a double-blind, randomized, placebo-controlled design, we tested 45 (18 female) healthy volunteers during a baseline session, after application of losartan or placebo (drug session), and during a follow-up session. During each session, participants engaged in a task in which they had to predict the probability of an electrical stimulation on every trial while the true shock contingencies switched repeatedly between phases of high and low shock threat. Computational reinforcement learning models were used to investigate learning dynamics. Results: Acute administration of losartan significantly reduced participants’ adjustment during both low-to-high and high-to-low threat changes. This was driven by reduced aversive learning rates in the losartan group during the drug session compared with baseline. The 50-mg drug dose did not induce reduction of blood pressure or change in reaction times, ruling out a general reduction in attention and engagement. Decreased adjustment of aversive expectations was maintained at a follow-up session 24 hours later. Conclusions: This study shows that losartan acutely reduces Pavlovian learning in aversive environments, thereby highlighting a potential role of the renin-angiotensin system in anxiety development

Gender Differences in Symptoms and Care Delivery for Chronic Obstructive Pulmonary Disease

Abstract Background: Morbidity and mortality for women with chronic obstructive pulmonary disease (COPD) are increasing, and little is known about gender differences in perception of COPD care. Methods: Surveys were administered to a convenience sample of COPD patients to evaluate perceptions about symptoms, barriers to care, and sources of information about COPD. Results: Data on 295 female and 273 male participants were analyzed. With similar frequencies, women and men reported dyspnea and rated their health as poor/very poor. Although more women than men reported annual household income <$30,000, no significant gender differences in frequency of health insurance, physician visits, or ever having had spirometry were detected. In adjusted models (1) women were more likely to report COPD diagnostic delay (odds ratio [OR] 1.66, 95% confidence interval [CI] 1.13-2.45, p=0.01), although anxiety (OR 1.83, 95% CI 1.10-3.06, p=0.02) and history of exacerbations (OR 1.60, 95% CI 1.08-2.37, p=0.01) were also significant predictors, (2) female gender was associated with difficulty reaching one's physician (OR 2.54, 95% CI 1.33-4.86, p=0.004), as was prior history of exacerbations (OR 2.25, 95% CI 1.21-4.20, p=0.01), and (3) female gender (OR 2.15, 95% CI 1.10-4.21, p=0.02) was the only significant predictor for finding time spent with their physician as insufficient. Conclusions: Significant gender-related differences in the perception of COPD healthcare delivery exist, revealing an opportunity to better understand what influences these attitudes and to improve care for both men and women.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98466/1/jwh%2E2012%2E3650.pd

Comparative effectiveness of intensity modulated radiation therapy to 3-dimensional conformal radiation in locally advanced lung cancer: pathological and clinical outcomes.

OBJECTIVE: Intensity-modulated radiotherapy (IMRT) has better normal-tissue sparing compared with 3-dimensional conformal radiation (3DCRT). We sought to assess the impact of radiation technique on pathological and clinical outcomes in locally advanced non-small cell lung cancer (LANSCLC) treated with a trimodality strategy. METHODS: Retrospective review of LANSCLC patients treated from August 2012 to August 2018 at Sheba Medical Center, Israel. The trimodality strategy consisted of concomitant chemoradiation to 60 Gray (Gy) followed by completion surgery. The planning target volume (PTV) was defined by co-registered PET/CT. Here we compare the pathological regression, surgical margin status, local control rates (LC), disease free (DFS) and overall survival (OS) between 3DCRT and IMRT. RESULTS: Our cohort consisted of 74 patients with mean age 62.9 years, male in 51/74 (69%), adenocarcinoma in 46/74 (62.1%), stage 3 in 59/74 (79.7%) and chemotherapy in 72/74 (97.3%). Radiation mean dose: 59.2 Gy (SD ± 3.8). Radiation technique : 3DCRT in 51/74 (68.9%), IMRT in 23/74 (31%). Other variables were similar between groups.Major pathological response (including pathological complete response or less than 10% residual tumor cells) was similar: 32/51 (62.7%) in 3DCRT and 15/23 (65.2%) in IMRT, p=0.83. Pathological complete response (pCR) rates were similar: 17/51 (33.3%) in 3DCRT and 8/23 (34.8%) in IMRT, p=0.9. Surgical margins were negative in 46/51 (90.1%) in 3DCRT vs. 17/19 (89.4%) in IMRT (p=1.0).The 2-year LC rates were 81.6% (95% CI 69-89.4%); DFS 58.3% (95% CI 45.5-69%) and 3-year OS 70% (95% CI57-80%). Comparing radiation techniques, there were no significant differences in LC (p=0.94), DFS (p=0.33) and OS (p=0.72). CONCLUSION: When used to treat LANSCLC in the neoadjuvant setting, both IMRT and 3DCRT produce comparable pathological and clinical outcomes. ADVANCES IN KNOWLEDGE: This study validates the real-world effectiveness of IMRT compared to 3DCRT

Genome-wide associations for birth weight and correlations with adult disease

Birth weight (BW) is influenced by both foetal and maternal factors and in observational studies is reproducibly associated with future risk of adult metabolic diseases including type 2 diabetes (T2D) and cardiovascular disease1. These lifecourse associations have often been attributed to the impact of an adverse early life environment. We performed a multi-ancestry genome-wide association study (GWAS) meta-analysis of BW in 153,781 individuals, identifying 60 loci where foetal genotype was associated with BW (P <5x10-8). Overall, ˜15% of variance in BW could be captured by assays of foetal genetic variation. Using genetic association alone, we found strong inverse genetic correlations between BW and systolic blood pressure (rg-0.22, P =5.5x10-13), T2D (rg-0.27, P =1.1x10-6) and coronary artery disease (rg-0.30, P =6.5x10-9) and, in large cohort data sets, demonstrated that genetic factors were the major contributor to the negative covariance between BW and future cardiometabolic risk. Pathway analyses indicated that the protein products of genes within BW-associated regions were enriched for diverse processes including insulin signalling, glucose homeostasis, glycogen biosynthesis and chromatin remodelling. There was also enrichment of associations with BW in known imprinted regions (P =1.9x10-4). We have demonstrated that lifecourse associations between early growth phenotypes and adult cardiometabolic disease are in part the result of shared genetic effects and have highlighted some of the pathways through which these causal genetic effects are mediated

Genome-wide associations for birth weight and correlations with adult disease

Birth weight (BW) has been shown to be influenced by both fetal and maternal factors and in observational studies is reproducibly associated with future risk of adult metabolic diseases including type 2 diabetes (T2D) and cardiovascular disease. These life-course associations have often been attributed to the impact of an adverse early life environment. Here, we performed a multi-ancestry genome-wide association study (GWAS) meta-analysis of BW in 153,781 individuals, identifying 60 loci where fetal genotype was associated with BW ($\textit{P}$  < 5 × 10$^{-8}$). Overall, approximately 15% of variance in BW was captured by assays of fetal genetic variation. Using genetic association alone, we found strong inverse genetic correlations between BW and systolic blood pressure ($\textit{R}$ $_{g}$ = -0.22, $\textit{P}$  = 5.5 × 10$^{-13}$), T2D ($\textit{R}$ $_{g}$ = -0.27, $\textit{P}$  = 1.1 × 10$^{-6}$) and coronary artery disease ($\textit{R}$ $_{g}$ = -0.30, $\textit{P}$  = 6.5 × 10$^{-9}$). In addition, using large -cohort datasets, we demonstrated that genetic factors were the major contributor to the negative covariance between BW and future cardiometabolic risk. Pathway analyses indicated that the protein products of genes within BW-associated regions were enriched for diverse processes including insulin signalling, glucose homeostasis, glycogen biosynthesis and chromatin remodelling. There was also enrichment of associations with BW in known imprinted regions ($\textit{P}$ = 1.9 × 10$^{-4}$). We demonstrate that life-course associations between early growth phenotypes and adult cardiometabolic disease are in part the result of shared genetic effects and identify some of the pathways through which these causal genetic effects are mediated.For a full list of the funders pelase visit the publisher's website and look at the supplemetary material provided. Some of the funders are: British Heart Foundation, Cancer Research UK, Medical Research Council, National Institutes of Health, Royal Society and Wellcome Trust

Maternal and fetal genetic effects on birth weight and their relevance to cardio-metabolic risk factors.

Birth weight variation is influenced by fetal and maternal genetic and non-genetic factors, and has been reproducibly associated with future cardio-metabolic health outcomes. In expanded genome-wide association analyses of own birth weight (n = 321,223) and offspring birth weight (n = 230,069 mothers), we identified 190 independent association signals (129 of which are novel). We used structural equation modeling to decompose the contributions of direct fetal and indirect maternal genetic effects, then applied Mendelian randomization to illuminate causal pathways. For example, both indirect maternal and direct fetal genetic effects drive the observational relationship between lower birth weight and higher later blood pressure: maternal blood pressure-raising alleles reduce offspring birth weight, but only direct fetal effects of these alleles, once inherited, increase later offspring blood pressure. Using maternal birth weight-lowering genotypes to proxy for an adverse intrauterine environment provided no evidence that it causally raises offspring blood pressure, indicating that the inverse birth weight-blood pressure association is attributable to genetic effects, and not to intrauterine programming.The Fenland Study is funded by the Medical Research Council (MC_U106179471) and Wellcome Trust