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NASA NTRS1993

A combustion products analyzer for contingency use during thermodegradation events on spacecraft

The Toxicology Laboratory at JSC and Exidyne Instrumentation Technologies (EIT) have developed a prototype Combustion Products Analyzer (CPA) to monitor, in real time, combustion products from a thermodegradation event on board spacecraft. The CPA monitors the four gases that are the most hazardous compounds (based on the toxicity potential and quantity produced) likely to be released during thermodegradation of synthetic materials: hydrogen fluoride (HF), hydrogen chloride (HCl), hydrogen cyanide (HCN), and carbon monoxide (CO). The levels of these compounds serve as markers to assist toxicologists in determining when the cabin atmosphere is safe for the crew to breathe following the contingency event. The CPA is a hand-held, battery-operated instrument containing four electrochemical sensors, one for each target gas, and a pump for drawing air across the sensors. The sensors are unique in their small size and zero-g compatibility. The immobilized electrolytes in each sensor permit the instrument to function in space and eliminate the possibility of electrolye leaks. The sample inlet system is equipped with a particulate filter that prevents clogging from airborne particulate matter. The CPA has a large digital display for gas concentrations and warming signals for low flow and low battery conditions. The CPA has flown on 13 missions beginning with STS 41 in Oct. 1990. Current efforts include the development of a microprocessor, an improved carbon monoxide sensor, and a ground-based test program to evaluate the CPA during actual thermodegradation of selected materials.

Wilson, Steve↗

NASA NTRS2003

Evaluation of Treadmill Exercise in a Lower Body Negative Pressure Chamber as a Countermeasure for Weightlessness-Induced Bone Loss: a Bed Rest Study with Identical Twins

Counteracting bone loss is required for future space exploration. We evaluated the ability of treadmill exercise in a LBNP chamber to counteract bone loss in a 30-day bed rest study. Eight pairs of identical twins were randomly assigned to sedentary control or exercise groups. Exercise within LBNP decreased the bone resorption caused by bed rest and may provide a countermeasure for spaceflight. INTRODUCTION: Bone loss is one of the greatest physiological challenges for extended-duration space missions. The ability of exercise to counteract weightlessness-induced bone loss has been studied extensively, but to date, it has proven ineffective. We evaluated the effectiveness of a combination of two countermeasures-treadmill exercise while inside a lower body negative pressure (LBNP) chamber-on bone loss during a 30-day bed rest study. MATERIALS AND METHODS: Eight pairs of identical twins were randomized into sedentary (SED) or exercise/LBNP (EX/LBNP) groups. Blood and urine samples were collected before, several times during, and after the 30-day bed rest period. These samples were analyzed for markers of bone and calcium metabolism. Repeated measures ANOVA was used to determine statistical significance. Because identical twins were used, both time and group were treated as repeated variables. RESULTS: Markers of bone resorption were increased during bed rest in samples from sedentary subjects, including the collagen cross-links and serum and urinary calcium concentrations. For N-telopeptide and deoxypyridinoline, there were significant (p < 0.05) interactions between group (SED versus EX/LBNP) and phase of the study (sample collection point). Pyridinium cross-links were increased above pre-bed rest levels in both groups, but the EX/LBNP group had a smaller increase than the SED group. Markers of bone formation were unchanged by bed rest in both groups. CONCLUSIONS: These data show that this weight-bearing exercise combined with LBNP ameliorates some of the negative effects of simulated weightlessness on bone metabolism. This protocol may pave the way to counteracting bone loss during spaceflight and may provide valuable information about normal and abnormal bone physiology here on Earth.

Twin Study↗

DOE OSTI2016

Identifying Urinary and Serum Exosome Biomarkers for Radiation Exposure Using a Data Dependent Acquisition and SWATH-MS Combined Workflow

Purpose: Early and accurate assessment of radiation injury by radiation-responsive biomarkers is critical for triage and early intervention. Biofluids such as urine and serum are convenient for such analysis. Recent research has also suggested that exosomes are a reliable source of biomarkers in disease progression. In the present study, we analyzed total urine proteome and exosomes isolated from urine or serum for potential biomarkers of acute and persistent radiation injury in mice exposed to lethal whole body irradiation (WBI). Methods and Materials: For feasibility studies, the mice were irradiated at 10.4 Gy WBI, and urine and serum samples were collected 24 and 72 hours after irradiation. Exosomes were isolated and analyzed using liquid chromatography mass spectrometry/mass spectrometry-based workflow for radiation exposure signatures. A data dependent acquisition and SWATH-MS combined workflow approach was used to identify significantly exosome biomarkers indicative of acute or persistent radiation-induced responses. For the validation studies, mice were exposed to 3, 6, 8, or 10 Gy WBI, and samples were analyzed for comparison. Results: A comparison between total urine proteomics and urine exosome proteomics demonstrated that exosome proteomic analysis was superior in identifying radiation signatures. Feasibility studies identified 23 biomarkers from urine and 24 biomarkers from serum exosomes after WBI. Urinary exosome signatures identified different physiological parameters than the ones obtained in serum exosomes. Exosome signatures from urine indicated injury to the liver, gastrointestinal, and genitourinary tracts. In contrast, serum showed vascular injuries and acute inflammation in response to radiation. Selected urinary exosomal biomarkers also showed changes at lower radiation doses in validation studies. Conclusions: Exosome proteomics revealed radiation- and time-dependent protein signatures after WBI. A total of 47 differentially secreted proteins were identified in urinary and serum exosomes. Together, these data showed the feasibility of defining biomarkers that could elucidate tissue-associated and systemic response caused by high-dose ionizing radiation. This is the first report using an exosome proteomics approach to identify radiation signatures.

62 RADIOLOGY AND NUCLEAR MEDICINE↗

DOE OSTI2019

Miscanthus sinensis multi-location trial: phenotypic analysis, genome-wide association, and genomic prediction

This dataset contains genotypic and phenotypic data, R scripts, and the results of analysis pertaining to a multi-location field trial of Miscanthus sinensis . Genome-wide association and genomic prediction were performed for biomass yield and 14 yield-component traits across six field trial locations in Asia and North America, using 46,177 single-nucleotide polymorphism (SNP) markers mined from restriction site-associated DNA sequencing (RAD-seq) and 568 M. sinensis accessions. Genomic regions and candidate genes were identified that can be used for breeding improved varieties of M. sinensis , which in turn will be used to generate new M. xgiganteus clones for biomass.

genome-wide association studies (GWAS)↗

DOE OSTI2016

MO-FG-CAMPUS-JeP3-05: Evaluation of 4D CT-On-Rails Target Localization Methods for Free Breathing Liver Stereotactic Body Radiotherapy (SBRT)

Purpose: Liver SBRT patients unable to tolerate breath-hold for radiotherapy are treated free-breathing with image guidance. Target localization using 3D CBCT requires extra margins to accommodate the respiratory motion. The purpose of this study is to evaluate the accuracy and reproducibility of 4D CT-on-rails in target localization for free-breathing liver SBRT. Methods: A Siemens SOMATOM CT-on-Rails 4D with Anzai Pressure Belt system was used both as the simulation and the localization CT. Fiducial marker was placed close to the center of the target prior to the simulation. Amplitude based sorting was used in the scan. Eight or sixteen phases of reconstructed CT sets (depends on breathing pattern) can be sent to Velocity to create the maximum intensity projection (MIP) image set. Target ITV and fiducial ITV were drawn based on the MIP image. In patient localization, a 4D scan was taken with the same settings as the sim scan. Images were registered to match fiducial ITVs. Results: Ten liver cancer patients treated for 50Gy over 5 fractions, with amplitudes of breathing motion ranging from 4.3–14.5 mm, were analyzed in this study. Results show that the Intra & inter fraction variability in liver motion amplitude significantly less than the baseline inter-fraction shifts in liver position. 90% of amplitude change is less than 3 mm. The differences in the D99 and D95 GTV dose coverage between the 4D CT-on-Rails and the CBCT plan were small (within 5%) for all the selected cases. However, the average PTV volume by using the 4D CT-on-Rails is 37% less than the CBCT PTV volume. Conclusion: Simulation and Registration using 4D CT-on-Rails provides accurate target localization and is unaffected by larger breathing amplitudes as seen with 3D CBCT image registration. Localization with 4D CT-on-Rails can significantly reduce the PTV volume with sufficient tumor.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2016

flowVS: channel-specific variance stabilization in flow cytometry

Abstract Background Comparing phenotypes of heterogeneous cell populations from multiple biological conditions is at the heart of scientific discovery based on flow cytometry (FC). When the biological signal is measured by the average expression of a biomarker, standard statistical methods require that variance be approximately stabilized in populations to be compared. Since the mean and variance of a cell population are often correlated in fluorescence-based FC measurements, a preprocessing step is needed to stabilize the within-population variances. Results We present a variance-stabilization algorithm, called flowVS, that removes the mean-variance correlations from cell populations identified in each fluorescence channel. flowVS transforms each channel from all samples of a data set by the inverse hyperbolic sine (asinh) transformation. For each channel, the parameters of the transformation are optimally selected by Bartlett’s likelihood-ratio test so that the populations attain homogeneous variances. The optimum parameters are then used to transform the corresponding channels in every sample. flowVS is therefore an explicit variance-stabilization method that stabilizes within-population variances in each channel by evaluating the homoskedasticity of clusters with a likelihood-ratio test. With two publicly available datasets, we show that flowVS removes the mean-variance dependence from raw FC data and makes the within-population variance relatively homogeneous. We demonstrate that alternative transformation techniques such as flowTrans, flowScape, logicle, and FCSTrans might not stabilize variance. Besides flow cytometry, flowVS can also be applied to stabilize variance in microarray data. With a publicly available data set we demonstrate that flowVS performs as well as the VSN software, a state-of-the-art approach developed for microarrays. Conclusions The homogeneity of variance in cell populations across FC samples is desirable when extracting features uniformly and comparing cell populations with different levels of marker expressions. The newly developed flowVS algorithm solves the variance-stabilization problem in FC and microarrays by optimally transforming data with the help of Bartlett’s likelihood-ratio test. On two publicly available FC datasets, flowVS stabilizes within-population variances more evenly than the available transformation and normalization techniques. flowVS-based variance stabilization can help in performing comparison and alignment of phenotypically identical cell populations across different samples. flowVS and the datasets used in this paper are publicly available in Bioconductor.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2016

Deciphering Natural Allelic Variation in Switchgrass for Biomass Yield and Quality Using a Nested Association Mapping Population

Switchgrass (Panicum virgatum L.) is a C4 grass with high biomass yield potential and a model species for bioenergy feedstock development. Understanding the genetic basis of quantitative traits is essential to facilitate genome-enabled breeding programs. The nested association mapping (NAM) analysis combines the best features of both bi-parental and association analyses and can provide high power and high resolution in QTL detection and will ensure significant improvements in biomass yield and quality. To develop a NAM population of switchgrass, 15 highly diverse genotypes with specific characteristics were selected from a diversity panel and crossed to a recurrent parent, AP13, a genotype selected for whole genome sequencing and parent of a mapping population. Ten genotypes from each of the 15 F1 families were then chain crossed. Progenies form each family were randomly selected to develop the NAM population. The switchgrass NAM population consists of a total of 2000 genotypes from 15 families. All the progenies, founder parents, F1 parents (n=2350) were evaluated in replicated field trials at Ardmore, OK and Knoxville, TN. Phenotypic data on plant height, tillering ability, regrowth, flowering time, and biomass yield were collected. Dried biomass samples were also analyzed using prediction equations of NIRS at the Noble Foundation and for lignin content, S/G ratio, and sugar release characteristics at the NREL. Genomic shotgun sequencing of 15 switchgrass NAM founder parental genomes at JGI produced 28-66 Gb high-quality sequence data. Alignment of these sequences with the reference genome, AP13 (v3.0), revealed that up to 99% of the genomic sequences mapped to the reference genome. A total of 2,149 individuals from NAM populations were sequenced by exome capture and two sets of 15 SNP matrices (one for each family) were generated. QTL associated with important traits have been identified and verified in breeding populations. The QTL detected and their associated markers can be used in molecular breeding programs to facilitate development of improved switchgrass cultivars for biofuel production.

09 BIOMASS FUELS↗

DOE OSTI2021

Comprehensive analysis of differentially expressed mRNA and circRNA in Ankylosing spondylitis patients’ platelets

Highlights: • 4996 mRNAs and 2942 circRNAs were identified differentially expressed in platelets of AS patients. • Several platelet-derived immune mediators were upregulated to regulate the interaction between platelets and immune cells. • Two downregulated circRNA were identified, and the corresponding circRNA-miRNA-mRNA regulatory network was constructed. • mRNAs and circRNAs found herein could be used as diagnostic biomarkers or new therapeutic targets. Ankylosing spondylitis (AS) is a chronic inflammatory disease significantly decreasing the quality of life. Platelets play an important and active role in the development of AS. Accumulating evidence demonstrated platelets contain diverse RNA repository inherited from megakaryocytes or microvesicles. Platelet RNAs are dynamically affected by pathological conditions and could be used as diagnostic or prognostic biomarkers. However, the role of the platelet RNAs in AS is elusive. In this study, we compared mRNA and circRNA profiles in platelets between AS patients and healthy controls using RNA sequencing and bioinformatic analysis, and found 4996 mRNAs and 2942 circRNAs were differently expressed. The significantly over-expressed mRNAs in AS patients are involved in platelet activity, gap junction, focal adhesion, rap1 and toll and Imd signaling pathway. The previous identified platelet-derived immune mediators such as P2Y1, P2Y12, PF4, GPIbα, CD40L, ICAM2, CCL5 (RANTES), TGF-β (TGF-β1 and TGF-β2) and PDGF (PDGFB and PDGFA) are also included in these over expressed mRNAs, implying these factors may trigger inflammatory cascades and promote the development of AS. Additionally, we found two down-regulated circRNA (circPTPN22 and circFCHSD2) from the intersection analyses of platelets and spinal ligament tissues of AS patients. The circRNA-miRNA-mRNA regulatory network of these two circRNAs was constructed, and the target mRNAs were enriched in Th17 cell differentiation, inflammatory bowel disease, cell adhesion molecules, cytokine-cytokine receptor interaction, Jak-STAT and Wnt signaling pathway, all these pathways participate in the bone remodeling and pro/anti-inflammatory immune regulation in AS. Then, qRT-PCR was performed to validate the expression of selected key mRNAs and circRNAs and the results demonstrated that the expression levels of P2Y12, GPIbα, circPTPN22 and circFCHSD2 were consistent with the sequencing analysis. In addition, the high expression of five predicted miRNAs interacting with circPTPN22 and circFCHSD2 were also detected in AS by qRT-PCR. Taken together, our study presents a comprehensive overview of mRNAs and circRNAs in platelets in AS patients and offers new insight into the mechanisms of platelet involving in the pathogenesis of AS. The mRNAs and circRNAs identified in this study may serve as candidates for diagnosis and targeted treatment of AS.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2021

A wearable patch for continuous analysis of thermoregulatory sweat at rest

The body naturally and continuously secretes sweat for thermoregulation during sedentary and routine activities at rates that can reflect underlying health conditions, including nerve damage, autonomic and metabolic disorders, and chronic stress. However, low secretion rates and evaporation pose challenges for collecting resting thermoregulatory sweat for non-invasive analysis of body physiology. Here we present wearable patches for continuous sweat monitoring at rest, using microfluidics to combat evaporation and enable selective monitoring of secretion rate. We integrate hydrophilic fillers for rapid sweat uptake into the sensing channel, reducing required sweat accumulation time towards real-time measurement. Along with sweat rate sensors, we integrate electrochemical sensors for pH, Cl - , and levodopa monitoring. We demonstrate patch functionality for dynamic sweat analysis related to routine activities, stress events, hypoglycemia-induced sweating, and Parkinson’s disease. By enabling sweat analysis compatible with sedentary, routine, and daily activities, these patches enable continuous, autonomous monitoring of body physiology at rest.

47 OTHER INSTRUMENTATION↗

DOE OSTI2022

Giant kelp genetic monitoring before and after disturbance reveals stable genetic diversity in Southern California

Given the impacts of climate change and other anthropogenic stressors on marine systems, there is a need to accurately predict how species respond to changing environments and disturbance regimes. The use of genetic tools to monitor temporal trends in populations gives ecologists the ability to estimate changes in genetic diversity and effective population size that may be undetectable by traditional census methods. Although multiple studies have used temporal genetic analysis, they usually involve commercially important species, and rarely sample before and after disturbance. In this study, we run a temporal analysis of giant kelp, Macrocystis pyrifera , genetic diversity over the scope of 10 years (2008-2018) using the same microsatellite marker panel to assess the genetic consequences of disturbance in several populations of giant kelp ( Macrocystis pyrifera ) in the Southern California Bight. The study is a rare pre- and post-disturbance microsatellite analysis that included declines to giant kelp caused by the 2015/16 El Nino Southern Oscillation event. We used canopy biomass estimated by remote sensing (Landsat) to quantify the extent of disturbance to kelp beds, and sea surface temperature data to understand how kelp was pushed towards its temperature limits during this period. Despite prolonged periods with decreased canopy at several sites, no changes in genetic structure and allelic richness were observed. We argue that giant kelp in the region is best described as a “patchy population” system where true extinctions are rare. We discuss how deep refugia of subsurface sporophytes and cryptic microscopic life stages could have kept genetic diversity through disturbance. Given the increasing effects of climate change and uncertainty in modeling impacts of species with cryptic life history stages, we suggest further investigation to reveal the role such stages play in species resilience. Genetic monitoring studies of sites selected by remote census demographic and climate surveys should be continued in the future given the predicted impacts of climate change.

Klingbeil, III, William H.↗

DOE OSTI2019

A proteome-wide assessment of the oxidative stress paradigm for metal and metal-oxide nanomaterials in human macrophages

Responsible implementation of engineered nanomaterials (ENMs) into commercial applications is an important societal issue, driving demand for new approaches for rapid and comprehensive evaluation of their bioactivity and safety. An essential part of any research focused on identifying potential hazard of ENMs is the appropriate selection of biological endpoints to evaluate. Herein, we use a tiered strategy employing both targeted biological assays and untargeted quantitative proteomics to elucidate the biological responses of human macrophages across a library of metal/metal oxide ENMs, raised as priority ENMs for investigation by NIEHS’s Nanomaterial Health Implications Research (NHIR) program. Our results show that quantitative cellular proteome profiles readily distinguish ENM types based on their cytotoxic potential according to induction of biological processes and pathways involved in the cellular antioxidant response, TCA cycle, oxidative stress, endoplasmic reticulum stress, and immune responses as major processes impacted. Interestingly, pathway analysis of differentially abundant proteins also revealed new cell processes that were influenced by all ENMs independent of cytotoxic potential. These included pathways previously implicated as cellular adaptive mechanisms to low levels of oxidative stress, including cell adhesion, protein translation and protein targeting pathways. Unsupervised clustering of all data revealed the most striking proteome changes that differentiated ENM classes involved a small subset of proteins involved in the oxidative stess response (HMOX1), protein chaperone functions (HS71B, DNJB1), and autophagy (SQSTM), providing a potential new panel of markers of ENM-induced cell stress. To our knowledge, the results represent the most comprehensive profiling of the biological responses to a library of ENMs conducted using quantitative mass spectrometry-based proteomics. The results provide a basis to identify the patterns of a diverse set of cellular pathways and cell processes impacted by ENM exposure in an important immune cell type, laying the foundation for multivariate, pathway-level structure activity assessments of ENMs in the future.

36 MATERIALS SCIENCE↗

DOE OSTI2019

In silico identification of key genes and signaling pathways targeted by a panel of signature microRNAs in prostate cancer

Accumulating evidence have suggested that some microRNAs are aberrantly expressed in prostate cancer. In our previous work, we had identified a panel of four differentially expressed microRNAs in prostate cancer. In the present study, we have investigated common molecular targets of this panel of miRNAs (DEMs) and key hub genes that can serve as potential candidate biomarkers in the pathogenesis and progression of prostate cancer. A joint bioinformatics approach was employed to identify differentially expressed genes (DEGs) in prostate cancer. Gene enrichment analysis followed by the protein–protein interaction (PPI) network construction and selection of hub genes was further performed using String and Cytoscape, respectively. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the identified hub genes was conducted using the Database for Annotation, Visualization and Integrated Discovery (DAVID) tool. In total, 496 genes were identified to be common targets of DEMs in prostate cancer and 13 key hub genes were identified from three modules of the PPI network of the DEGs. Further top five genes viz Rhoa, PI3KCA, CDC42, MAPK3, TP53 were used for Enrichment analysis which revealed their association with vital cellular and functional pathways in prostate cancer indicating their potential as candidate biomarkers in prostate cancer.

62 RADIOLOGY AND NUCLEAR MEDICINE↗

DOE OSTI2021

PCGF3 promotes the proliferation and migration of non-small cell lung cancer cells via the PI3K/AKT signaling pathway

Highlights: • PCGF3 is highly expressed in non-small cell lung cancer (NSCLC) tissues. • PCGF3 overexpression positively correlates with lymph node metastasis and p-TNM staging. • PCGF3 promotes NSCLC cells proliferation by regulating CyclinB1, CyclinD1, and CDK4. • PCGF3 promotes NSCLC cells migration by regulating RhoA, RhoC, and CDC42. • PCGF3 promotes malignant behavior in NSCLC by positively regulating the PI3K/AKT signaling pathway. The Polycomb Group Ring Finger 3 (PCGF3) protein has been reported to be significantly upregulated in pancreatic islet tumors and related to signal transduction; however, its detailed mechanisms and biological roles in other tumors, including non-small cell lung cancer (NSCLC), remain unclear. This study investigated the function of PCGF3 in NSCLC and further elucidated its mechanism of action. The immunohistochemical analysis of 86 selected lung cancer tissues revealed that PCGF3 was highly expressed in NSCLC tissues and positively correlated with lymph node metastasis and p-TNM staging. Additionally, PCGF3 promoted cell proliferation in lung cancer by regulating CyclinB1, CyclinD1, and CDK4 expression, and also promoting their migration by regulating RhoA, RhoC, and CDC42. Furthermore, PCGF3 affected both the proliferation and migration of lung cancer cells by regulating the PI3K/AKT pathway, as verified by inhibiting this pathway using LY294002. The findings of this study suggested that PCGF3 is associated with poor prognosis in patients with NSCLC and could therefore be an important biomarker for treating and preventing NSCLC.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Differential Mobility Spectrometry-Mass Spectrometry (DMS-MS) in Radiation Biodosimetry: Rapid and High-Throughput Quantitation of Multiple Radiation Biomarkers in Nonhuman Primate Urine

High-throughput methods to assess radiation exposure are a priority due to concerns that include nuclear power accidents, the spread of nuclear weapon capability, and the risk of terrorist attacks. Metabolomics, the assessment of small molecules in an easily accessible sample, is the most recent method to be applied for the identification of biomarkers of the biological radiation response with a useful dose-response profile. Profiling for biomarker identification is frequently done using an LC-MS platform which has limited throughput due to the time-consuming nature of chromatography. We present here a chromatography-free simplified method for quantitative analysis of seven metabolites in urine with radiation dose-response using urine samples provided from the Pannkuk et al. (2015) study of long-term (7-day) radiation response in nonhuman primates (NHP). The stable isotope dilution (SID) analytical method consists of sample preparation by strong cation exchange-solid phase extraction (SCX-SPE) to remove interferences and concentrate the metabolites of interest, followed by differential mobility spectrometry (DMS) ion filtration to select the ion of interest and reduce chemical background, followed by mass spectrometry (overall SID-SPE-DMS-MS). Since no chromatography is used, calibration curves were prepared rapidly, in under 2 h (including SPE) for six simultaneously analyzed radiation biomarkers. The seventh, creatinine, was measured separately after 2500× dilution. Creatinine plays a dual role, measuring kidney glomerular filtration rate (GFR), and indicating kidney damage at high doses. The current quantitative method using SID-SPE-DMS-MS provides throughput which is 7.5 to 30 times higher than that of LC-MS and provides a path to pre-clinical radiation dose estimation. .

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

DOE OSTI2014

Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

60 APPLIED LIFE SCIENCES↗

DOE OSTI2018

Senescence chips for ultrahigh-throughput isolation and removal of senescent cells

Cellular senescence plays an important role in organismal aging and age-related diseases. However, it is challenging to isolate low numbers of senescent cells from small volumes of biofluids for downstream analysis. Furthermore, there is no technology that could selectively remove senescent cells in a high-throughput manner. In this work, we developed a novel microfluidic chip platform, termed senescence chip, for ultrahigh-throughput isolation and removal of senescent cells. The core component of our senescence chip is a slanted and tunable 3D micropillar array with a variety of shutters in the vertical direction for rapid cell sieving, taking advantage of the characteristic cell size increase during cellular senescence. The 3D configuration achieves high throughput, high recovery rate, and device robustness with minimum clogging. We demonstrated proof-of-principle applications in isolation and enumeration of senescent mesenchymal stem cells (MSCs) from undiluted human whole blood, and senescent cells from mouse bone marrow after total body irradiation, with the single cell resolution. After scale-up to a multilayer and multichannel structure, our senescence chip achieved ultrahigh-throughput removal of senescent cells from human whole blood with an efficiency of over 70% at a flow rate of 300 ml/hr. Sensitivity and specificity of our senescence chips could be augmented with implementation of multiscale size separation, and identification of background white blood cells using their cell surface markers such as CD45. With the advantages of high throughput, robustness, and simplicity, our senescence chips may find wide applications and contribute to diagnosis and therapeutic targeting of cellular senescence.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS2008

Lunar Dust Effects on Spacesuit Systems: Insights from the Apollo Spacesuits

Systems and components of selected Apollo A7L/A7LB flight-article spacesuits that were worn on the lunar surface have been studied to determine the degree to which they suffered contamination, abrasion and wear or loss of function due to effects from lunar soil particles. Filter materials from the lithium hydroxide (LiOH) canisters from the Apollo Command Module were also studied to determine the amount and type of any lunar dust particles they may have captured from the spacecraft atmosphere. The specific spacesuit study materials include the outermost soft fabric layers on Apollo 12 and 17 integrated thermal micrometeorite garment assemblies and outermost fabrics on Apollo 17 extravehicular pressure gloves. In addition, the degree of surface wear in the sealed wrist rotation bearing from Apollo 16 extravehicular and intravehicular pressure gloves was evaluated and compared. Scanning electron microscope examination of the Apollo 12 T-164 woven TeflonO fabric confirms the presence of lunar soil particles and the ability of these particles to cause separation and fraying of the Teflon fibers. Optical imaging, chemical analysis and particle sampling applied to the outer fabric of the Apollo 17 spacesuit has identified Ti as a potentially useful chemical marker for comparing the amount of lunar soil retained on different areas of the spacesuit outer fabric. High-yield particle sampling from the Apollo 17 fabric surfaces using adhesive tape found 80% of particles on the fabric are lunar soil particles averaging 10.5 m in diameter, with the rest being intrinsic fabric materials or environmental contaminants. Analysis of the mineralogical composition of the lunar particles found that on a grain-count basis the particle population is dominated by plagioclase feldspar and various types of glassy particles derived mostly from soil agglutinates, with a subordinate amount of pyroxene. On a grain size basis, however, the pyroxene grains are generally a factor of 2 larger than glass and plagioclase, so conversion of the data to a modal (volume %) basis results in pyroxene becoming the modally dominant particle type with glass and plagioclase significantly less abundant. When comparisons are made to the modal composition of lunar soil at the Apollo 17 landing site, the results suggest that pyroxene particles have overall better retention on the spacesuit outer fabric compared to plagioclase and especially glass. Scanning electron microscopy revealed no measureable difference in the amount of wear and abrasion in the wrist rotation bearing of an Apollo 16 pressure glove worn only in the spacecraft and one worn only for extravehicular activity on the lunar surface. The results suggest either that the bearing prevented entry of lunar dust, or that dust was not sufficiently abrasive to damage the bearing, or both.

Christoffersen, Roy↗

DOE OSTI2016

SU-F-T-20: Novel Catheter Lumen Recognition Algorithm for Rapid Digitization

Purpose: Manual catheter recognition remains a time-consuming aspect of high-dose-rate brachytherapy (HDR) treatment planning. In this work, a novel catheter lumen recognition algorithm was created for accurate and rapid digitization. Methods: MatLab v8.5 was used to create the catheter recognition algorithm. Initially, the algorithm searches the patient CT dataset using an intensity based k-means filter designed to locate catheters. Once the catheters have been located, seed points are manually selected to initialize digitization of each catheter. From each seed point, the algorithm searches locally in order to automatically digitize the remaining catheter. This digitization is accomplished by finding pixels with similar image curvature and divergence parameters compared to the seed pixel. Newly digitized pixels are treated as new seed positions, and hessian image analysis is used to direct the algorithm toward neighboring catheter pixels, and to make the algorithm insensitive to adjacent catheters that are unresolvable on CT, air pockets, and high Z artifacts. The algorithm was tested using 11 HDR treatment plans, including the Syed template, tandem and ovoid applicator, and multi-catheter lung brachytherapy. Digitization error was calculated by comparing manually determined catheter positions to those determined by the algorithm. Results: he digitization error was 0.23 mm ± 0.14 mm axially and 0.62 mm ± 0.13 mm longitudinally at the tip. The time of digitization, following initial seed placement was less than 1 second per catheter. The maximum total time required to digitize all tested applicators was 4 minutes (Syed template with 15 needles). Conclusion: This algorithm successfully digitizes HDR catheters for a variety of applicators with or without CT markers. The minimal axial error demonstrates the accuracy of the algorithm, and its insensitivity to image artifacts and challenging catheter positioning. Future work to automatically place initial seed positions would improve the algorithm speed.

60 APPLIED LIFE SCIENCES↗