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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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7 records
All sourcesNASA NTRSDOE OSTI
NASA NTRS2006

Three-Dimensional Human Bronchial-Tracheal Epithelial Tissue-Like Assemblies (TLAs) as Hosts for Severe Acute Respiratory Syndrome (SARS)-CoV Infection

A three-dimensional (3-D) tissue-like assembly (TLA) of human bronchial-tracheal mesenchymal (HBTC) cells with an overlay of human bronchial epithelial (BEAS-2B) cells was constructed using a NASA Bioreactor to survey the infectivity of SARS-CoV. This TLA was inoculated with a low passage number Urbani strain of SARS-CoV. At selected intervals over a 10-day period, media and cell aliquots of the 3-D TLA were harvested for viral titer assay and for light and electron microscopy examination. All viral titer assays were negative in both BEAS-2B two-dimensional monolayer and TLA. Light microscopy immunohistochemistry demonstrated antigen-antibody reactivity with anti-SARS-CoV polyclonal antibody to spike and nuclear proteins on cell membranes and cytoplasm. Coronavirus Group 2 cross-reactivity was demonstrated by positive reaction to anti-FIPV 1 and anti-FIPV 1 and 2 antibodies. TLA examination by transmission electron microscopy indicated increasing cytoplasmic vacuolation with numerous electron-dense bodies measuring 45 to 270 nm from days 4 through 10. There was no evidence of membrane blebbing, membrane duplication, or fragmentation of organelles in the TLAs. However, progressive disruption of endoplasmic reticulum was observed throughout the cells. Antibody response to SARS-CoV specific spike and nucleocapsid glycoproteins, cross-reactivity with FIPV antibodies, and the cytoplasmic pathology suggests this HBTE TLA model is permissive to SARS-CoV infection.

Suderman, M. T.↗
NASA NTRS2015

New Insights into Amino Acid Preservation in the Early Oceans Using Modern Analytical Techniques

Protein- and non-protein-amino acids likely occupied the oceans at the time of the origin and evolution of life. Primordial soup-, hydrothermal vent-, and meteoritic-processes likely contributed to this early chemical inventory. Prebiotic synthesis and carbonaceous meteorite studies suggest that non-protein amino acids were likely more abundant than their protein-counterparts. Amino acid preservation before abiotic and biotic destruction is key to biomarker availability in paleoenvironments and remains an important uncertainty. To constrain primitive amino acid lifetimes, a 1992 archived seawater/beach sand mixture was spiked with D,L-alanine, D,L-valine (Val), alpha-aminoisobutyric acid (alpha-AIB), D,L-isovaline (Iva), and glycine (Gly). Analysis by high performance liquid chromatography with fluorescence detection (HPLC-FD) showed that only D-Val and non-protein amino acids were abundant after 2250 days. The mixture was re-analyzed in 2012 using HPLC-FD and a triple quadrupole mass spectrometer (QqQ-MS). The analytical results 20 years after the inception of the experiment were strikingly similar to those after 2250 days. To confirm that viable microorganisms were still present, the mixture was re-spiked with Gly in 2012. Aliquots were collected immediately after spiking, and at 5- and 9-month intervals thereafter. Final HPLC-FD/QqQ-MS analyses were performed in 2014. The 2014 analyses revealed that only alpha-AIB, D,L-Iva, and D-Val remained abundant. The disappearance of Gly indicated that microorganisms still lived in the mixture and were capable of consuming protein amino acids. These findings demonstrate that non-protein amino acids are minimally impacted by biological degradation and thus have very long lifetimes under these conditions. Primitive non-protein amino acids from terrestrial synthesis, or meteorite in-fall, likely experienced great-er preservation than protein amino acids in paleo-oceanic environments. Such robust molecules may have reached a steady state concentration dependent on ocean circulation through hydrothermal systems and synthetic input processes. We are presently trying to estimate this concentration.

Parker, Eric T.↗
NASA NTRSDate not supplied

Method Development for In-situ Detection of Latent Herpesvirus DNA from Saliva using Nanopore Sequencing

Research toward latent herpesvirus reactivation has been intensively addressed through Space Shuttle and International Space Station (ISS) investigations. This work has provided the understanding that persistent reactivation of herpesviruses from asymptomatic crew can be detected through viral shedding in saliva, urine, and blood. Occasionally, viral reactivation from the latency stage can pose a threat to crew health (clinical manifestation) before, during, and after flight missions. Furthermore, previous work detailing correlations with immunity indicate that monitoring viral reactivation could be implemented to assess potential immune dysfunction. While in-flight monitoring is desirable, there is no well-established procedure or method for real-time evaluations, and research to date has relied on postflight, ground-based analysis. The development of portable molecular technologies like the miniPCR™ (miniPCR Bio) thermal cycler and the MinION™ sequencer (Oxford Nanopore Technologies) have confirmed that real-time monitoring is possible in extreme and low resource environments. These devices, combined with simple sample preparation methods, have been used to demonstrate bacterial identification onboard the ISS, as well as rapid viral detection in remote locations on Earth. The work here builds upon previous molecular advancements onboard the ISS toward the development and validation of a spaceflight-compatible method for viral detection from crew samples. Several herpesviruses can be detected in saliva, which provides a non-invasive means to collect samples for monitoring. While the basis for this method lies in previous spaceflight investigations, key points for method optimization include DNA extraction from saliva, viral primer selection, and bioinformatic processes for data analysis. To increase viral yield, numerous DNA extraction methods have been evaluated and will be discussed in detail. For initial development and testing, varicella-zoster virus (VZV) is being targeted though open reading frame 51 and 63 (ORF51, ORF63), as the replication origin-binding protein is highly expressed during latency. Optimization of the thermal cycling parameters has resulted in the ability to test the entire process. The full method has been tested with both viral VZV DNA standards and saliva spiked with varying concentrations of VZV. Viral sequence data were mapped to the reference sequence using minimap2. Prior to mapping, DNA sequencing reads were filtered for length and quality, barcodes were removed, and alignment identity calculated. Following further assessments, statistics were compared across multiple sequencing experiments and are being used to determine the success of the protocols. Forward work will include the incorporation of herpes simplex virus 1 (HSV-1) and Epstein-Barr virus (EBV) primers as well as the validation of results to the terrestrial qPCR standard assay. Upon full validation of the developed method, saliva will be collected from 20 healthy subjects and spiked with viral DNA. These samples will be split and assayed with the MinION and standard qPCR assay.

Hang N Nguyen↗
NASA NTRS2000

Skeletal muscle Ca(2+)-independent kinase activity increases during either hypertrophy or running

Spikes in free Ca(2+) initiate contractions in skeletal muscle cells, but whether and how they might signal to transcription factors in skeletal muscles of living animals is unknown. Since previous studies in non-muscle cells have shown that serum response factor (SRF) protein, a transcription factor, is phosphorylated rapidly by Ca(2+)/calmodulin (CaM)-dependent protein kinase after rises in intracellular Ca(2+), we measured enzymatic activity that phosphorylates SRF (designated SRF kinase activity). Homogenates from 7-day-hypertrophied anterior latissimus dorsi muscles of roosters had more Ca(2+)-independent SRF kinase activity than their respective control muscles. However, no differences were noted in Ca(2+)/CaM-dependent SRF kinase activity between control and trained muscles. To determine whether the Ca(2+)-independent and Ca(2+)/CaM-dependent forms of Ca(2+)/CaM-dependent protein kinase II (CaMKII) might contribute to some of the SRF kinase activity, autocamtide-3, a synthetic substrate that is specific for CaMKII, was employed. While the Ca(2+)-independent form of CaMKII was increased, like the Ca(2+)-independent form of SRF kinase, no alteration in CaMKII occurred at 7 days of stretch overload. These observations suggest that some of SRF phosphorylation by skeletal muscle extracts could be due to CaMKII. To determine whether this adaptation was specific to the exercise type (i.e., hypertrophy), similar measurements were made in the white vastus lateralis muscle of rats that had completed 2 wk of voluntary running. Although Ca(2+)-independent SRF kinase was increased, no alteration occurred in Ca(2+)/CaM-dependent SRF kinase activity. Thus any role of Ca(2+)-independent SRF kinase signaling has downstream modulators specific to the exercise phenotype.

NASA Program Fundamental Space Biology↗
NASA NTRS1983

Rain-induced spring wheat harvest losses

When rain or a combination of rain and high humidity delay wheat harvest, losses can occur in grain yield and/or grain quality. Yield losses can result from shattering, from reduction in test weight, and in the case of windrowed grain, from rooting of sprouting grain at the soil: windrow contact. Losses in grain quality can result from reduction in test weight and from sprouting. Sprouting causes a degradation of grain proteins and starches, hence flour quality is reduced, and the grain price deteriorates to the value of feed grain. Although losses in grain yield and quality are rain-induced, these losses do not necessarily occur because a standing or windrowed crop is wetted by rain. Spike water concentration in hard red spring wheat must be increased to about 45-49% before sprouting is initiated in grain that has overcome dormancy. The time required to overcome this dormancy after the cultivar has dried to 12 to 14% water concentration differs with hard red spring cultivars. The effect of rain on threshing-ready standing and windrowed hard red spring wheat grain yeild and quality was evaluated. A goal was to develop the capability to forecast the extent of expected loss of grain yield and quality from specific climatic events that delay threshing.

Bauer, A.↗
NASA NTRS2003

Detection of Strand Cleavage And Oxidation Damage Using Model DNA Molecules Captured in a Nanoscale Pore

We use a biological nano-scale pore to distinguish among individual DNA hairpins that differ by a single site of oxidation or a nick in the sugar-phosphate backbone. In earlier work we showed that the protein ion channel alpha-hemolysin can be used as a detector to distinguish single-stranded from double-stranded DNA, single base pair and single nucleotide differences. This resolution is in part a result of sensitivity to structural changes that influence the molecular dynamics of nucleotides within DNA. The strand cleavage products we examined here included a 5-base-pair (5-bp) hairpin with a 5-prime five-nucleotide overhang, and a complementary five-nucleotide oligomer. These produced predictable shoulder-spike and rapid near-full blockade signatures, respectively. When combined, strand annealing was monitored in real time. The residual current level dropped to a lower discrete level in the shoulder-spike blockade signatures, and the duration lengthened. However, these blockade signatures had a shorter duration than the unmodified l0bp hairpin. To test the pore sensitivity to nucleotide oxidation, we examined a 9-bp hairpin with a terminal 8-oxo-deoxyguanosine (8-oxo-dG), or a penultimate 8-oxo-dG. Each produced blockade signatures that differed from the otherwise identical control 9bp hairpins. This study showed that DNA structure is modified sufficiently by strand cleavage or oxidation damage at a single site to alter in a predictable manner the ionic current blockade signatures produced. This technique improves the ability to assess damage to DNA, and can provide a simple means to help characterize the risks of radiation exposure. It may also provide a method to test radiation protection.

Vercoutere, W.↗
NASA NTRS2003

Discrimination of Single Base Pair Differences Among Individual DNA Molecules Using a Nanopore

The protein toxin alpha-hemolysin form nanometer scale channels across lipid membranes. Our lab uses a single channel in an artificial lipid bilayer in a patch clamp device to capture and examine individual DNA molecules. This nanopore detector used with a support vector machine (SVM) can analyze DNA hairpin molecules on the millisecond time scale. We distinguish duplex stem length, base pair mismatches, loop length, and single base pair differences. The residual current fluxes also reveal structural molecular dynamics elements. DNA end-fraying (terminal base pair dissociation) can be observed as near full blockades, or spikes, in current. This technique can be used to investigate other biological processes dependent on DNA end-fraying, such as the processing of HIV DNA by HIV integrase.

Vercoutere, Wenonah↗