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At least 343 records · Page 19

Simulating photospheric Doppler velocity fields

A method is described for constructing artificial data that realistically simulate photospheric velocity fields. The velocity fields include rotation, differential rotation, meridional circulation, giant cell convection, supergranulation, convective limb shift, p-mode oscillations, and observer motion. Data constructed by this method can be used for testing algorithms designed to extract and analyze these velocity fields in real Doppler velocity data.

Hathaway, David H.↗

Regulation of Osteoblast Survival by the Extracellular Matrix and Gravity

Spaceflight adversely affects the skeleton, posing a substantial risk to astronaut's health during long duration missions. The reduced bone mass observed in growing animals following spaceflight is due at least in part to inadequate bone formation by osteoblasts. Thus, it is of central importance to identify basic cellular mechanisms underlying normal bone formation. The fundamental ideas underlying our research are that interactions between extracellular matrix proteins, integrin adhesion receptors, cytoplasmic signaling and cytoskeletal proteins are key ingredients for the proper functioning of osteoblasts, and that gravity impacts these interactions. As an in vitro model system we used primary fetal rat calvarial cells which faithfully recapitulate osteoblast differentiation characteristically observed in vivo. We showed that specific integrin receptors ((alpha)3(beta)1), ((alpha)5(beta)1), ((alpha)8(betal)1) and extracellular matrix proteins (fibronectin, laminin) were needed for the differentiation of immature osteoblasts. In the course of maturation, cultured osteoblasts switched from depending on fibronectin and laminin for differentiation to depending on these proteins for their very survival. Furthermore, we found that manipulating the gravity vector using ground-based models resulted in activation of key intracellular survival signals generated by integrin/extracellular matrix interactions. We are currently testing the in vivo relevance of some of these observations using targeted transgenic technology. In conclusion, mechanical factors including gravity may participate in regulating survival via cellular interactions with the extracellular matrix. This leads us to speculate that microgravity adversely affects the survival of osteoblasts and contributes to spaceflight-induced osteoporosis.

Globus. Ruth K.↗

Neuregulin: First Steps Towards a Structure

Neuregulins are growth factor domain proteins with diverse bioactivities, such as cell proliferation, receptor binding, and differentiation. Neureguh- 1 binds to two members of the ErbB class I tyrosine kinase receptors, ErbB3 and ErbB4. A number of human cancers overexpress the ErbB receptors, and neuregulin can modulate the growth of certain cancer types. Neuregulin-1 has been shown to promote the migration of invasive gliomas of the central nervous system. Neuregulin has also been implicated in schizophrenia, multiple sclerosis and abortive cardiac abnormalities. The full function of neuregulin-1 is not known. In this study we are inserting a cDNA clone obtained from American Type Culture Collection into E.coli expression vectors to express neuregulin- 1 protein. Metal chelate affinity chromatography is used for recombinant protein purification. Crystallization screening will proceed for X-ray diffraction studies following expression, optimization, and protein purification. In spite of medical and scholarly interest in the neuregulins, there are currently no high-resolution structures available for these proteins. Here we present the first steps toward attaining a high-resolution structure of neuregulin- 1, which will help enable us to better understand its function

Ferree, D. S.↗

ATP as a biomarker of viable microorganisms in clean-room facilities

A new firefly luciferase bioluminescence assay method that differentiates free extracellular ATP (dead cells, etc.) from intracellular ATP (viable microbes) was used to determine the viable microbial cleanliness of various clean-room facilities. For comparison, samples were taken from both clean-rooms, where the air was filtered to remove particles >0.5 microm, and ordinary rooms with unfiltered air. The intracellular ATP was determined after enzymatically degrading the sample's free ATP. Also for comparison, cultivable microbial populations were counted on nutrient-rich trypticase soy agar (TSA) plates. Both the cultivable and ATP-based determinations indicate that the microbial burden was lower in clean-room facilities than in ordinary rooms. However, there was no direct correlation between the two sets of measurements because the two assays measured very different populations. A large fraction of the samples yielded no colony formers on TSA, but were positive for intracellular ATP. Subsequently, genomic DNA was isolated directly from selected samples and 16S rDNA fragments were cloned and sequenced, identifying nearest neighbors, many of which are known to be noncultivable in the media employed. It was concluded that viable microbial contamination can be reliably monitored by measurement of intracellular ATP, and that this method may be considered superior to cultivable colony counts due to its speed and its ability to report the presence of viable but noncultivable organisms. When the detection of nonviable microbes is of interest, the ATP assay can be supplemented with DNA analysis.

NASA Discipline Environmental Health↗

Osteoblast fibronectin mRNA, protein synthesis, and matrix are unchanged after exposure to microgravity

The well-defined osteoblast line, MC3T3-E1 was used to examine fibronectin (FN) mRNA levels, protein synthesis, and extracellular FN matrix accumulation after growth activation in spaceflight. These osteoblasts produce FN extracellular matrix (ECM) known to regulate adhesion, differentiation, and function in adherent cells. Changes in bone ECM and osteoblast cell shape occur in spaceflight. To determine whether altered FN matrix is a factor in causing these changes in spaceflight, quiescent osteoblasts were launched into microgravity and were then sera activated with and without a 1-gravity field. Synthesis of FN mRNA, protein, and matrix were measured after activation in microgravity. FN mRNA synthesis is significantly reduced in microgravity (0-G) when compared to ground (GR) osteoblasts flown in a centrifuge simulating earth's gravity (1-G) field 2.5 h after activation. However, 27.5 h after activation there were no significant differences in mRNA synthesis. A small but significant reduction of FN protein was found in the 0-G samples 2.5 h after activation. Total FN protein 27.5 h after activation showed no significant difference between any of the gravity conditions, however, there was a fourfold increase in absolute amount of protein synthesized during the incubation. Using immunofluorescence, we found no significant differences in the amount or in the orientation of the FN matrix after 27.5 h in microgravity. These results demonstrate that FN is made by sera-activated osteoblasts even during exposure to microgravity. These data also suggest that after a total period of 43 h of spaceflight FN transcription, translation, or altered matrix assembly is not responsible for the altered cell shape or altered matrix formation of osteoblasts.

Flight Experiment↗

Optical remote measurement of toxic gases

Enactment of the Clean Air Act Amendments (CAAA) of 1990 has resulted in increased ambient air monitoring needs for industry, some of which may be met efficiently using open-path optical remote sensing techniques. These techniques include Fourier transform spectroscopy, differential optical absorption spectroscopy, laser long-path absorption, differential absorption lidar, and gas cell correlation spectroscopy. With this regulatory impetus, it is an opportune time to consider applying these technologies to the remote and/or path-averaged measurement and monitoring of toxic gases covered by the CAAA. This article reviews the optical remote sensing technology and literature for that application.

Review↗

Studies of mineralization in tissue culture: optimal conditions for cartilage calcification

The optimal conditions for obtaining a calcified cartilage matrix approximating that which exists in situ were established in a differentiating chick limb bud mesenchymal cell culture system. Using cells from stage 21-24 embryos in a micro-mass culture, at an optimal density of 0.5 million cells/20 microliters spot, the deposition of small crystals of hydroxyapatite on a collagenous matrix and matrix vesicles was detected by day 21 using X-ray diffraction, FT-IR microscopy, and electron microscopy. Optimal media, containing 1.1 mM Ca, 4 mM P, 25 micrograms/ml vitamin C, 0.3 mg/ml glutamine, no Hepes buffer, and 10% fetal bovine serum, produced matrix resembling the calcifying cartilage matrix of fetal chick long bones. Interestingly, higher concentrations of fetal bovine serum had an inhibitory effect on calcification. The cartilage phenotype was confirmed based on the cellular expression of cartilage collagen and proteoglycan mRNAs, the presence of type II and type X collagen, and cartilage type proteoglycan at the light microscopic level, and the presence of chondrocytes and matrix vesicles at the EM level. The system is proposed as a model for evaluating the events in cell mediated cartilage calcification.

Non-NASA Center↗

Technology Advancement for Active Remote Sensing of Carbon Dioxide from Space Using the ASCENDS CarbonHawk Experiment Simulator: First Results

The ASCENDS CarbonHawk Experiment Simulator (ACES) is a newly developed lidar developed at NASA Langley Research Center and funded by NASA's Earth Science Technology Office (ESTO) Instrument Incubator Program (IIP) that seeks to advance technologies critical to measuring atmospheric column carbon dioxide (CO2) mixing ratios in support of the NASA Active Sensing of CO2 Emissions over Nights, Days, and Seasons (ASCENDS) mission. The technology advancements targeted include: (1) increasing the power-aperture product to approach ASCENDS mission requirements by implementing multi-aperture telescopes and multiple co-aligned laser transmitters; (2) incorporating high-efficiency, high-power Erbium-Doped Fiber Amplifiers (EDFAs); (3) developing and incorporating a high-bandwidth, low-noise HgCdTe detector and transimpedence amplifier (TIA) subsystem capable of long-duration autonomous operation on Global Hawk aircraft, and (4) advancing algorithms for cloud and aerosol discrimination. The ACES instrument architecture is being developed for operation on high-altitude aircraft and will be directly scalable to meet the ASCENDS mission requirements. These technologies are critical towards developing not only spaceborne instruments but also their airborne simulators, with lower platform requirements for size, mass, and power, and with improved instrument performance for the ASCENDS mission. ACES transmits five laser beams: three from commercial EDFAs operating near 1.57 microns, and two from the Exelis oxygen (O2) Raman fiber laser amplifier system operating near 1.26 microns. The three EDFAs are capable of transmitting up to 10 watts average optical output power each and are seeded by compact, low noise, stable, narrow-linewidth laser sources stabilized with respect to a CO2 absorption line using a multi-pass gas absorption cell. The Integrated-Path Differential Absorption (IPDA) lidar approach is used at both wavelengths to independently measure the CO2 and O2 column number densities and retrieve the average column CO2 mixing ratio. The ACES receiver uses three fiber-coupled 17.8-cm diameter athermal telescopes. The transmitter assembly consists of five fiber-coupled laser collimators and an associated Risley prism pair for each laser to co-align the outgoing laser beams and to align them with the telescope field of view. The backscattered return signals collected by the three telescopes are combined in a fiber bundle and sent to a single low noise detector. The detector/TIA development has improved the existing detector subsystem by increasing its bandwidth to 4.7 MHz from 500 kHz and increasing the duration of autonomous, service-free operation periods from 4 hours to >24 hours. The new detector subsystem enables the utilization of higher laser modulation rates, which provides greater flexibility for implementing advanced thin-cloud discrimination algorithms as well as improving range-determination resolution and error reduction. The cloud/aerosol discrimination algorithm development by Langley and Exelis features a new suite of algorithms for the minimization/elimination of bias errors in the return signal induced by the presence of intervening thin clouds. Multiple laser modulation schemes are being tested in an effort to significantly mitigate the effects of thin clouds on the retrieved CO2 column amounts. Full instrument development concluded in the spring of 2014. After ground range tests of the instrument, ACES successfully completed six test flights on the Langley Hu-25 aircraft in July, 2014, and recorded data at multiple altitudes over land and ocean surfaces with and without intervening clouds. Preliminary results from these test flights will be presented in this paper.

Obland, Michael D.↗

Evaluating Electroluminescence Imaging and Image Processing as a Quantitative Solar Cell Characterization Method

Mitigating dust accumulation on the surface of solar arrays is crucial for maintaining maximum power output. We propose investigating electroluminescence imaging paired with image processing as a means of evaluating various dust mitigation techniques. Image processing was able to clearly differentiate between pristine and dusted solar cells. Paired with traditional analysis techniques, this method proves to be a quick and powerful characterization tool.

Photovoltaic↗

In vitro chick pre-cardiac explant tissue differentiation during spaceflight on SpaceHab-02

Chick precardiac tissue explants were cultured on the 8-day mission of STS-60, space shuttle Discovery. Development of in vitro cultures of precardiac chick tissue from embryo stages 5 though 8 (H-H) were initiated during orbit and were terminated after approximately fifteen hours of 37 degree C culture. Transmission electron microscopy and tritiated thymidine studies were performed postflight. No significant differences in cell proliferation were observed between flight and ground controls. Electron-microscopic studies revealed stage 8 explants were capable of differentiation during flight in a pattern which matched ground control tissues. As anticipated, stage 7 explant tissues had differentiated to a lesser extent compared to stage 8 tissues. Interestingly, stage 7 precardiac explant flight tissue differentiation was less than ground control tissue. This difference in differentiation between flight and ground cultures was enhanced in stage 6 tissues, as high levels of myofibril organization were only seen in ground controls. Other cellular components such as Golgi apparatus, junctional complexes, and mitochondria were present and appeared normal and healthy.

NASA Program NSCORT↗

Light-driven solute transport in Halobacterium halobium

The cell membrane of Halobacterium halobium exhibits differential regions which contain crystalline arrays of a single kind of protein, termed bacteriorhodopsin. This bacterial retinal-protein complex resembles the visual pigment and, after the absorption of protons, translocates H(+) across the cell membrane, leading to an electrochemical gradient for protons between the inside and the outside of the cell. Thus, light is an alternate source of energy in these bacteria, in addition to terminal oxidation. The paper deals with work on light-driven transport in H. halobium with cell envelope vesicles. The discussion covers light-driven movements of H(+), Na(+), and K(+); light-driven amino acid transport; and apparent allosteric control of amino acid transport. The scheme of energy coupling in H. halobium vesicles appears simple, its quantitative details are quite complex and reveal regulatory phenomena. More knowledge is required of the way the coupling components are regulated by the ion gradients present.

Lanyi, J. K.↗

Differential RNA regulation by staphylococcal enterotoxins A and B in murine macrophages

Staphylococcal enterotoxin A (SEA) is significantly better than enterotoxin B (SEB) in activating tumor necrosis factor (TNF) secretion by B6MP102 cells. Both toxins bound to B6MP102 cells; however, SEB competed less effectively with SEA than SEA competed with SEB. This suggested that receptors unique to SEA were present on B6MP102 cells. Signal transduction occurred in response to both toxins. Within 30 s after addition, SEA and SEB significantly increased the F-actin concentration in B6MP102 cells. However, only SEA induced increased TNF mRNA levels. B6MP102 cells incubated with interferon-gamma and SEB secreted TNF. However, enhanced mRNA expression was delayed and the concentration of TNF secreted was less than that of B6MP102 cells stimulated with SEA. Although these data suggest that receptors unique to SEA are present on B6MP102 cells, they also indicate that staphylococcal enterotoxins differentially regulate TNF at the RNA level, perhaps because of differences in binding to the plasma membrane.

NASA Discipline Cell Biology↗

Laser measurement of extinction coefficients of highly absorbing liquids

A coaxial dual-channel laser system has been developed for the measurement of extinction coefficients of highly absorbing liquids. An empty wedge-shaped sample cell is first translated laterally through a He-Ne laser beam to measure the differential thickness using interference fringes in reflection. The wedge cell is carefully filled with the oil sample and translated through the coaxially positioned dye laser beam for the differential attenuation or extinction measurement. Optional use of the instrumentation as a single-channel extinction measurement system and also as a refractometer is detailed. The system and calibration techniques were applied to the measurement of two crude oils whose extinction values were required to complete the analysis of airborne laser data gathered over four controlled spills.

Hoge, F. E.↗

A Novel Bone Marrow Single Cell Atlas of Mechanotransduction in Microgravity, Normal Gravity, Exercise, and Hindlimb Unloading for WT and CDKN1A-Null Regenerative Mice

Mechanical loading of adult stem cell progenitors is a key factor in modulating their proliferation, differentiation, and tissue regenerative potential, with loading generally promoting tissue formation and unloading mediating tissue loss. To address the role of mechanotransduction in maintaining stem cell-based tissue regenerative health, we generated a single cell transcriptomic atlas mapping responses of femur bone marrow mesenchymal and hematopoietic lineages to a range of altered mechanical loading conditions, both in WT B1629SF2/J mice as well as the p21/CDKN1A-null regenerative mice. We selected the femur marrow compartment as a model because of the diversity and high numbers of stem cell progenitor stages present, and because it undergoes static and cyclic hydrostatic pressure loading associated with weight-bearing and ambulation. Our study included normally loaded mice at 1g, unloading in microgravity during spaceflight as well as tail suspension hindlimb unloading, and voluntary running wheel exercise. Basal, one-year natural aging, and habitat controls were also conducted. Overall, the atlas encompasses 18 different experimental conditions with N=3 mice per condition and includes more than 500,000 single cell expressomes. Key specific findings include: increased mature reticulocyte populations in aging and unloading mice compared to active mice; greater hematopoietic and mesenchymal differentiating progenitors cluster identification in CDKN1A-null samples, especially the exercise model; distinct pseudotime cell trajectory shifts in the hematopoietic lineage for spaceflight and unloaded mice; and shifts in monocytic cell populations toward osteoclastic bone degenerative lineages in unloading and spaceflight samples. Overall mechanical loading shows increased marrow progenitor population differentiation while unloading is associated with increased CDKN1A expression and maintenance of marrow population stemness. Deletion of CDKN1A appears to remove a negative check on progenitor lineages commitment to differentiation. Finally, the Bone Marrow Mechanotransduction Single Cell Atlas will serve as a reference tool for studying marrow regenerative responses across a range of mechanical environments as they relate to CDKN1A status.

single-cell↗

Electrophoresis and isoelectric focusing of whole cell and membrane proteins from the extremely halophilic archaebacteria

The subunits from two purified halobacterial membrane enzymes (ATPase and nitrate reductase) behaved differently with respect to isoelectric focusing, silver staining and interaction with ampholytes. Differential behavior was also observed in whole cell proteins from Halobacterium saccharovorum regarding resolution in two-dimensional gels and silver staining. It is proposed that these differences reflect the existence of two classes of halobacterial proteins.

Stan-Lotter, Helga↗

Improved Cell Culture Method for Growing Contracting Skeletal Muscle Models

An improved method for culturing immature muscle cells (myoblasts) into a mature skeletal muscle overcomes some of the notable limitations of prior culture methods. The development of the method is a major advance in tissue engineering in that, for the first time, a cell-based model spontaneously fuses and differentiates into masses of highly aligned, contracting myotubes. This method enables (1) the construction of improved two-dimensional (monolayer) skeletal muscle test beds; (2) development of contracting three-dimensional tissue models; and (3) improved transplantable tissues for biomedical and regenerative medicine applications. With adaptation, this method also offers potential application for production of other tissue types (i.e., bone and cardiac) from corresponding precursor cells.

Marquette, Michele L.↗

Vector-averaged gravity-induced changes in cell signaling and vitamin D receptor activity in MG-63 cells are reversed by a 1,25-(OH)2D3 analog, EB1089

Skeletal unloading in an animal hindlimb suspension model and microgravity experienced by astronauts or as a result of prolonged bed rest causes site-specific losses in bone mineral density of 1%-2% per month. This is accompanied by reductions in circulating levels of 1,25-(OH)(2)D(3), the active metabolite of vitamin D. 1,25-(OH)(2)D(3), the ligand for the vitamin D receptor (VDR), is important for calcium absorption and plays a role in differentiation of osteoblasts and osteoclasts. To examine the responses of cells to activators of the VDR in a simulated microgravity environment, we used slow-turning lateral vessels (STLVs) in a rotating cell culture system. We found that, similar to cells grown in microgravity, MG-63 cells grown in the STLVs produce less osteocalcin, alkaline phosphatase, and collagen Ialpha1 mRNA and are less responsive to 1,25-(OH)(2)D(3). In addition, expression of VDR was reduced. Moreover, growth in the STLV caused activation of the stress-activated protein kinase pathway (SAPK), a kinase that inhibits VDR activity. In contrast, the 1,25-(OH)(2)D(3) analog, EB1089, was able to compensate for some of the STLV-associated responses by reducing SAPK activity, elevating VDR levels, and increasing expression of osteocalcin and alkaline phosphatase. These studies suggest that, not only does simulated microgravity reduce differentiation of MG-63 cells, but the activity of the VDR, an important regulator of bone metabolism, is reduced. Use of potent, less calcemic analogs of 1,25-(OH)(2)D(3) may aid in overcoming this defect. Copyright 2002 Elsevier Science Inc.

Foton Project↗

Three-Dimensional Cell Culture Models for Infectious Disease and Drug Development

Three-dimensional (3-D) cell cultures hold enormous potential to advance our understanding of infectious disease and to effectively translate basic cellular research into clinical applications. Using novel NASA bioreactor technology, the rotating wall vessel (RWV), we have engineered physiologically relevant 3-D human tissue culture models for infectious disease studies. The design of the RWV is based on the understanding that organs and tissues function in a 3-D environment, and that this 3-D architecture is critical for the differentiated form and function of tissues in vivo. The RWV provides large numbers of cells which are amenable to a wide variety of experimental manipulations and provides an easy, reproducible, and cost-effective approach to enhance differentiated features of cell culture models.

Nickerson, Cheryl A.↗