Data Exploration, Quality Control and Testing in Single-Cell qPCR-Based Gene Expression Experiments

Cell populations are never truly homogeneous; individual cells exist in biochemical states that define functional differences between them. New technology based on microfluidic arrays combined with multiplexed quantitative polymerase chain reactions (qPCR) now enables high-throughput single-cell gene expression measurement, allowing assessment of cellular heterogeneity. However very little analytic tools have been developed specifically for the statistical and analytical challenges of single-cell qPCR data. We present a statistical framework for the exploration, quality control, and analysis of single-cell gene expression data from microfluidic arrays. We assess accuracy and within-sample heterogeneity of single-cell expression and develop quality control criteria to filter unreliable cell measurements. We propose a statistical model accounting for the fact that genes at the single-cell level can be on (and for which a continuous expression measure is recorded) or dichotomously off (and the recorded expression is zero). Based on this model, we derive a combined likelihood-ratio test for differential expression that incorporates both the discrete and continuous components. Using an experiment that examines treatment-specific changes in expression, we show that this combined test is more powerful than either the continuous or dichotomous component in isolation, or a t-test on the zero-inflated data. While developed for measurements from a specific platform (Fluidigm), these tools are generalizable to other multi-parametric measures over large numbers of events.Comment: 9 pages, 5 figure

Impact of +Gz TO -Gz Induced Fluid Shifts on Ocular and Cerebral Parameters During Simulated Orion Re-Entry

No abstract availabl

Modelflow Estimates of Stroke Volume Do Not Correlate With Doppler Ultrasound Estimates During Upright Posture

Orthostatic intolerance affects 6080% of astronauts returning from longduration missions, representing a significant risk to completing missioncritical tasks. While likely multifactorial, a reduction in stroke volume (SV) represents one factor contributing to orthostatic intolerance during stand and head up tilt (HUT) tests. Current measures of SV during stand or HUT tests use Doppler ultrasound and require a trained operator and specialized equipment, restricting its use in the field. BeatScope (Finapres Medical Systems BV, The Netherlands) uses a modelflow algorithm to estimate SV from continuous blood pressure waveforms in supine subjects; however, evidence supporting the use of Modelflow to estimate SV in subjects completing stand or HUT tests remain scarce. Furthermore, because the blood pressure device is held extended at heart level during HUT tests, but allowed to rest at the side during stand tests, changes in the finger arterial pressure waveform resulting from arm positioning could alter modelflow estimated SV. The purpose of this project was to compare Doppler ultrasound and BeatScope estimations of SV to determine if BeatScope can be used during stand or HUT tests. Finger photoplethysmography was used to acquire arterial pressure waveforms corrected for hydrostatic fingertoheart height using the Finometer (FM) and Portapres (PP) arterial pressure devices in 10 subjects (5 men and 5 women) during a stand test while simultaneous estimates of SV were collected using Doppler ultrasound. Measures were made after 5 minutes of supine rest and while subjects stood for 5 minutes. Next, SV estimates were reacquired while each arm was independently raised to heart level, a position similar to tilt testing. Supine SV estimates were not significantly different between all three devices (FM: 68+/-20, PP: 71+/-21, US: 73+/-21 ml/beat). Upon standing, the change in SV estimated by FM (18+/-8 ml) was not different from PP (21+/-12), but both were significantly less than US (37+/-16 ml, p<.05). Raising finger BP devices to heart level caused no significant change in SV measured with any of the devices (FM: 1.5+/-19, PP: 1.7+/-26, US: 0.5+/-6), although variability was 36x greater as assessed by both blood pressure devices compared to US. Retrospective analysis of blood pressure data to assess SV in 11 supine subjects revealed significantly different estimates between methods (FM: 95+/-17, US: 75+/-32, p<.05), but the change in SV resulting from HUT was similar between methods (FM: 37+/-9, US: 40+/-18 ml). However, the correlation coefficient determined from pairs of SV estimated by US and FM was weak (r2=0.03). These data suggest Modelflow cannot be used in lieu of Doppler ultrasound to estimate SV during stand or HUT tests. Further investigation should focus on identifying factors contributing to differences between these measurement techniques in order to make use of a simple method for assessing beatbybeat changes in SV during postural changes, especially during field testing

High Intensity Resistive and Rowing Exercise Countermeasures Do Not Prevent Orthostatic Intolerance Following 70 Days of Bed Rest

More than 60% of US astronauts participating in Mir and early International Space Station missions (greater than 5 months) were unable to complete a 10min 80 deg headup tilt test on landing day. This high incidence of postspaceflight orthostatic intolerance may be related to limitations of the inflight exercise hardware that prevented high intensity training. PURPOSE: This study sought to determine if a countermeasure program that included intense lowerbody resistive and rowing exercises designed to prevent cardiovascular and musculoskeletal deconditioning during 70 days of 6 deg head-down tilt bed rest (BR), a spaceflight analog, also would protect against post BR orthostatic intolerance. METHODS: Sixteen males participated in this study and performed no exercise (Control, n=10) or performed an intense supine exercise protocol with resistive and aerobic components (Exercise, n=6). On 3 days/week, exercise subjects performed lower body resistive exercise and a 30min continuous bout of rowing (greater than or equal to 75% max heart rate). On 3 other days/week, subjects performed only highintensity, intervalstyle rowing. Orthostatic intolerance was assessed using a 15min 80 deg headup tilt test performed 2 days (BR2) before and on the last day of BR (BR70). Plasma volume was measured using a carbon monoxide rebreathing technique on BR3 and before rising on the first recovery day (BR+0). RESULTS: Following 70 days of BR, tilt tolerance time decreased significantly in both the Control (BR2: 15.0 +/- 0.0, BR70: 9.9 +/- 4.6 min, mean +/- SD) and Exercise (BR2: 12.2 +/- 4.7, BR70: 4.9 +/- 1.9 min) subjects, but the decreased tilt tolerance time was not different between groups (Control: 34 +/- 31, Exercise: 56 +/- 16%). Plasma volume also decreased (Control: 0.56 +/- 0.40, Exercise: 0.48 +/- 0.33 L) from pre to postBR, with no differences between groups (Control: 18 +/- 11%, Exerciser: 15 +/-1 0%). CONCLUSIONS: These findings confirm previous reports in shorter BR studies that the performance of an exercise countermeasure protocol by itself during BR does not prevent orthostatic intolerance or plasma volume loss. This suggests that protection against orthostatic intolerance in astronauts following longduration spaceflight will require an additional intervention, such as periodic orthostatic stress, fluid repletion, and/or lowerbody compression garments

Autonomous Diagnostic Imaging Performed by Untrained Operator Using Augmented Reality as a Form of "Just-in-Time" Training

We will address the Human Factors and Performance Team, "Risk of performance errors due to training deficiencies" by improving the JIT training materials for ultrasound and OCT imaging by providing advanced guidance in a detailed, timely, and user-friendly manner. Specifically, we will (1) develop an audio-visual tutorial using AR that guides non-experts through an abdominal trauma ultrasound protocol; (2) develop an audio-visual tutorial using AR to guide an untrained operator through the acquisition of OCT images; (3) evaluate the quality of abdominal ultrasound and OCT images acquired by untrained operators using AR guidance compared to images acquired using traditional JIT techniques (laptop-based training conducted before image acquisition); and (4) compare the time required to complete imaging studies using AR tutorials with images acquired using current JIT practices to identify areas for time efficiency improvements. Two groups of subjects will be recruited to participate in this study. Operator-subjects, without previous experience in ultrasound or OCT, will be asked to perform both procedures using either the JIT training with AR technology or the traditional JIT training via laptop. Images acquired by inexperienced operator-subjects will be scored by experts in that imaging modality for diagnostic and research quality; experts will be blinded to the form of JIT used to acquire the images. Operator-subjects also will be asked to submit feedback to improve the training modules used during the scans to improve future training modules. Scanned-subjects will be a small group individuals from whom all images will be acquired

International Capital Flows and the Returns to Safe Assets in the United States, 2003-2007

Part of the International Finance Discussion Paper

Model Flow Determination of Stroke Volume

No abstract availabl

Florida\u27s Mystery Coral-Killer Identified

An unusual coral disease appeared on the Florida Reef Tract in June 1995. It was distinct in its microbiology, its pattern of tissue degradation, the species susceptible to it, and its regional distribution. Symptoms included a sharp line between healthy and diseased tissue, as occurs with other coral diseases, but the pathogen responsible for the new outbreak seemed more virulent, affected a wider variety of species, and destroyed tissue much more rapidly than these other \u27line\u27 or \u27band\u27 diseases. We have identified the pathogen responsible for this new disease as a new species of Sphingomonas

NASA's Functional Task Test: High Intensity Exercise Improves the Heart Rate Response to a Stand Test Following 70 Days of Bedrest

Cardiovascular adaptations due to spaceflight are modeled with 6deg head-down tilt bed rest (BR) and result in decreased orthostatic tolerance. We investigated if high-intensity resistive and aerobic exercise with and without testosterone supplementation would improve the heart rate (HR) response to a 3.5-min stand test and how quickly these changes recovered following BR. During 70 days of BR male subjects performed no exercise (Control, n=10), high intensity supine resistive and aerobic exercise (Exercise, n=9), or supine exercise plus supplemental testosterone (Exercise+T, n=8; 100 mg i.m., weekly in 2-week on/off cycles). We measured HR for 2 min while subjects were prone and for 3 min after standing twice before and 0, 1, 6, and 11 days after BR. Mixed-effects linear regression models were used to evaluate group, time, and interaction effects. Compared to pre-bed rest, prone HR was elevated on BR+0 and BR+1 in Control, but not Exercise or Exercise+T groups, and standing HR was greater in all 3 groups. The increase in prone and standing HR in Control subjects was greater than either Exercise or Exercise+T groups and all groups recovered by BR+6. The change in HR from prone to standing more than doubled on BR+0 in all groups, but was significantly less in the Exericse+T group compared to the Control, but not Exercise group. Exercise reduces, but does not prevent the increase in HR observed in response to standing. The significantly lower HR response in the Exercise+T group requires further investigation to determine physiologic significance