Identification of SNPs and INDELS in swine transcribed sequences using short oligonucleotide microarrays

<p>Abstract</p> <p>Background</p> <p>Genome-wide detection of single feature polymorphisms (SFP) in swine using transcriptome profiling of day 25 placental RNA by contrasting probe intensities from either Meishan or an occidental composite breed with Affymetrix porcine microarrays is presented. A linear mixed model analysis was used to identify significant breed-by-probe interactions.</p> <p>Results</p> <p>Gene specific linear mixed models were fit to each of the log₂transformed probe intensities on these arrays, using fixed effects for breed, probe, breed-by-probe interaction, and a random effect for array. After surveying the day 25 placental transcriptome, 857 probes with a q-value ≤ 0.05 and |fold change| ≥ 2 for the breed-by-probe interaction were identified as candidates containing SFP. To address the quality of the bioinformatics approach, universal pyrosequencing assays were designed from Affymetrix exemplar sequences to independently assess polymorphisms within a subset of probes for validation. Additionally probes were randomly selected for sequencing to determine an unbiased confirmation rate. In most cases, the 25-mer probe sequence printed on the microarray diverged from Meishan, not occidental crosses. This analysis was used to define a set of highly reliable predicted SFPs according to their probability scores.</p> <p>Conclusion</p> <p>By applying a SFP detection method to two mammalian breeds for the first time, we detected transition and transversion single nucleotide polymorphisms, as well as insertions/deletions which can be used to rapidly develop markers for genetic mapping and association analysis in species where high density genotyping platforms are otherwise unavailable.</p> <p>SNPs and INDELS discovered by this approach have been publicly deposited in NCBI's SNP repository dbSNP. This method is an attractive bioinformatics tool for uncovering breed-by-probe interactions, for rapidly identifying expressed SNPs, for investigating potential functional correlations between gene expression and breed polymorphisms, and is robust enough to be used on any Affymetrix gene expression platform.</p

Detection of transcriptional difference of porcine imprinted genes using different microarray platforms

BACKGROUND: Presently, multiple options exist for conducting gene expression profiling studies in swine. In order to determine the performance of some of the existing microarrays, Affymetrix Porcine, Affymetrix Human U133+2.0, and the U.S. Pig Genome Coordination Program spotted glass oligonucleotide microarrays were compared for their reproducibility, coverage, platform independent and dependent sensitivity using fibroblast cell lines derived from control and parthenogenic porcine embryos. RESULTS: Array group correlations between technical replicates demonstrated comparable reproducibility in both Affymetrix arrays. Glass oligonucleotide arrays showed greater variability and, in addition, approximately 10% of probes had to be discarded due to slide printing defects. Probe level analysis of Affymetrix Human arrays revealed significant variability within probe sets due to the effects of cross-species hybridization. Affymetrix Porcine arrays identified the greatest number of differentially expressed genes amongst probes common to all arrays, a measure of platform sensitivity. Affymetrix Porcine arrays also identified the greatest number of differentially expressed known imprinted genes using all probes on each array, an ad hoc measure of realistic performance for this particular experiment. CONCLUSION: We conclude that of the platforms currently available and tested, the Affymetrix Porcine array is the most sensitive and reproducible microarray for swine genomic studies

Communication and proximity effects on outcomes attributable to sense of presence in distance bioinformatics education

<p>Abstract</p> <p>Background</p> <p>Online learning is increasingly popular in medical education and sense of presence has been posited as a factor contributing to its success. Communication media influences on sense of presence and learning outcomes were explored in this study. Test performance and ratings of instruction and technology, factors influenced by sense of presence, are compared under four conditions involving different media and degrees of student physical presence: 1) videoconference co-located, 2) webcast co-located, 3) videoconference dispersed, and 4) webcast dispersed.</p> <p>Methods</p> <p>Eighty one first to forth year medical students heard a lecture on telemedicine and were asked to collaboratively search a telemedicine website under conditions where the lecture was delivered by videoconference or one way streaming (webcast) and where students were either co-located or dispersed. In the videoconference conditions, co-located students could use the technology to interact with the instructor and could interact with each other face to face, while the dispersed students could use the technology to interact with both the instructor and each other. In the webcast conditions, all students could use chat to communicate with the instructor or each other, although the co-located students also could interact orally. After hearing the lecture, students collaboratively searched a telemedicine website, took a test on lecture-website content and rated the instruction and the technology they used. Test scores on lecture and website content and ratings of instruction and technology for the four conditions were compared with analysis of variance and chi-square tests.</p> <p>Results</p> <p>There were no significant differences in overall measures, although there were on selected ratings of instruction. Students in both webcast conditions indicated they were encouraged more to follow up on their own and felt instruction was more interactive than co-located videoconferencing students. Dispersed videoconferencing students indicated the highest levels of interaction and there was evidence they interacted more.</p> <p>Conclusion</p> <p>Results do not strongly support proximity as a sense of presence factor affecting performance and attitudes, but do suggest communication medium may affect interactivity.</p

CMB-S4: Forecasting Constraints on Primordial Gravitational Waves

CMB-S4---the next-generation ground-based cosmic microwave background (CMB) experiment---is set to significantly advance the sensitivity of CMB measurements and enhance our understanding of the origin and evolution of the Universe, from the highest energies at the dawn of time through the growth of structure to the present day. Among the science cases pursued with CMB-S4, the quest for detecting primordial gravitational waves is a central driver of the experimental design. This work details the development of a forecasting framework that includes a power-spectrum-based semi-analytic projection tool, targeted explicitly towards optimizing constraints on the tensor-to-scalar ratio, $r$, in the presence of Galactic foregrounds and gravitational lensing of the CMB. This framework is unique in its direct use of information from the achieved performance of current Stage 2--3 CMB experiments to robustly forecast the science reach of upcoming CMB-polarization endeavors. The methodology allows for rapid iteration over experimental configurations and offers a flexible way to optimize the design of future experiments given a desired scientific goal. To form a closed-loop process, we couple this semi-analytic tool with map-based validation studies, which allow for the injection of additional complexity and verification of our forecasts with several independent analysis methods. We document multiple rounds of forecasts for CMB-S4 using this process and the resulting establishment of the current reference design of the primordial gravitational-wave component of the Stage-4 experiment, optimized to achieve our science goals of detecting primordial gravitational waves for $r > 0.003$ at greater than $5\sigma$, or, in the absence of a detection, of reaching an upper limit of $r < 0.001$ at $95\%$ CL.Comment: 24 pages, 8 figures, 9 tables, submitted to ApJ. arXiv admin note: text overlap with arXiv:1907.0447

CMB-S4

We describe the stage 4 cosmic microwave background ground-based experiment CMB-S4

CMB-S4: Forecasting Constraints on Primordial Gravitational Waves

Abstract: CMB-S4—the next-generation ground-based cosmic microwave background (CMB) experiment—is set to significantly advance the sensitivity of CMB measurements and enhance our understanding of the origin and evolution of the universe. Among the science cases pursued with CMB-S4, the quest for detecting primordial gravitational waves is a central driver of the experimental design. This work details the development of a forecasting framework that includes a power-spectrum-based semianalytic projection tool, targeted explicitly toward optimizing constraints on the tensor-to-scalar ratio, r, in the presence of Galactic foregrounds and gravitational lensing of the CMB. This framework is unique in its direct use of information from the achieved performance of current Stage 2–3 CMB experiments to robustly forecast the science reach of upcoming CMB-polarization endeavors. The methodology allows for rapid iteration over experimental configurations and offers a flexible way to optimize the design of future experiments, given a desired scientific goal. To form a closed-loop process, we couple this semianalytic tool with map-based validation studies, which allow for the injection of additional complexity and verification of our forecasts with several independent analysis methods. We document multiple rounds of forecasts for CMB-S4 using this process and the resulting establishment of the current reference design of the primordial gravitational-wave component of the Stage-4 experiment, optimized to achieve our science goals of detecting primordial gravitational waves for r > 0.003 at greater than 5σ, or in the absence of a detection, of reaching an upper limit of r < 0.001 at 95% CL

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

SELDI-TOF-MS Analysis of Transcriptional Activation Protein Binding to Response Elements Regulating Carcinogenesis Enzymes

Abstract: The risk from chemical carcinogens and environmental toxins is dependent on the metabolic balance between bioactivation and detoxification enzymes. Therefore, agents that alter enzyme expression are critical factors in toxicity. Enhancement or suppression of enzyme activities through gene expression is in part regulated by interactions between specific DNA promoter response elements and specific transcription proteins. DNA-protein interactions are dependent upon translocation of proteins from the cytoplasm to the nucleus and the affinity of proteins for binding to transcription promoter sequences. A key factor in both processes is the intracellular redox state, which influences protein-protein interactions and protein-DNA binding and can be altered by exposure to electrophiles, antioxidants and oxidative stress. Oxidative stress levels can be readily detected by measurable effects on the intracellular glutathione (GSH):glutathione disulfide redox potential, the major intracellularredox buffer. Alterations in the GSH redox pool can directly affect enzyme activity by altering disulfide bonds in the transcription factors regulating enzyme expression. These may affect: 1) specific DNA-protein and protein-protein interactions, 2) cyst(e)ine redox state within transcriptional proteins and 3) translocation of transcription proteins from cytoplasmic to nuclear compartments within the cell. The studies reported here are designed to investigate the relative changes in enzyme expression in response to cellular redox potential changes using the new proteomics technology of surface enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI). Treatment of HeLa and HT29 human cell lines to increase the expression of enzymes that are upregulated by oxidative stress was used as a model system to determine the efficacy of the SELDI technology in measuring changes in transcriptional protein binding to transcriptional response elements. An important goal is to determine whether the SELDI will allow simultaneous studies of multiple transcriptional protein-DNA interactions in response to controlled oxidative stress. This will provide a better understanding of the effect of electrophilic carcinogens and oxidants on the balance between activation and detoxification mechanisms in chemical carcinogenesis

Risk factors for postoperative hyperglycemia in neonates

OBJECTIVE Postoperative hyperglycemia has been shown to be associated with higher morbidity and mortality in pediatric patients. Data on risk factors for neonatal patients is limited. The objective of this study was to identify pre- and intraoperative risk factors associated with postoperative glucose in neonates. METHODS We conducted a retrospective cohort study of neonates after surgical procedures between January and December 2016 in a quaternary neonatal intensive care unit. The primary outcome was hyperglycemia defined as serum glucose ≥8.3 mmol/L during the first 4 hours postoperatively. Secondary outcomes included death and length of stay. We assessed the association of risk factors with the postoperative glucose. RESULTS In total, 206 surgical procedures (171 patients) were evaluated, among which 178 had serum glucose values during the first 4 hours postoperatively available. The incidence of hyperglycemia was 54% (n = 96). The median (IQR) glucose during the first 4 hours in NICU was 8.4 (6.52-10.65) mmol/L. Risk factors for postoperative hyperglycemia were intraoperative glucose infusion rate (GIR) and gestational age. There was a non-linear relationship between gestational age and postoperative hyperglycemia. Mortality occurred in 6 (7%) in the no-hyperglycemia group and 3 (3%) in the hyperglycemia group (p = 0.31). CONCLUSIONS Among the risk factors, intraoperative GIR was identified as a modifiable factor that can reduce postoperative hyperglycemia. A non-linear relationship of gestational age with postoperative glucose provides new insights that may help advance our understanding of the complex mechanisms of glucose homeostasis in neonates