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At least 19 records

Solar Array (Radiated/Non-Radiated): Materials Characterization and Cryogenic Thermal Cycling Qualification for the Planned Europa Clipper Mission (ECM)

Two special instruments were custom built for our critical study and manufactured and employed to test the SA materials of interest to cryogenic temperature conditions. We have procured custom manufactured two independent systems such as Thermo -Mechanical Analyzer (TMA) with Three Point Bend Probe (TPBP) module and Dilatometer. TMA can be used to measure Young’ s modulus or elastic modulus (e -modulus). Dilatometer can be used to measure Co -efficient of Thermal Expansion (CTE). We have optimized the materials configuration to measure physical properties of radiated and non -radiated materials for CTE and e -modulus, which will be useful to assess materials and processes reliability for cryogenic temperature applications. We have successfully implemented for some materials that are of interest to solar panels for the planned ECM project to measure CTE and Young's modulus. Also a separate study was undertaken to thermal cycle qualify SAs for ECM project. One can do thermal cycling to temperatures of 50 K and 133 K using liquid Helium (Lq He). This is in reality exorbitantly too expensive and is also inefficient. Furthermore, this is not technically controllable test to 133 K and 50 K temperatures. This is also too expensive to accomplish. Therefore, a vacuum set -up was made along with a cryostat successfully. This is a closed loop system where we do not lose the Helium gas. We were able to get to 35 K for a given test coupon size. These temperatures are lower than what we would intend to qualify the solar array technologies. This will allow to have some margin at cold temperatures. We successfully completed the qualification of solar array technologies down to 50 K temperatures. This is the first time we are reporting this accomplishment. In this paper, we will present the solar array materials properties for cryogenic temperatures down to 20 K temperatures and the thermal cycling qualification test results from 133 K to 50 K for three times the mission life of 120 thermal cycles to meet JPL Design.

Ramesham, Rajeshuni

Ectomycorrhizal Community Structure and Soil Characteristics of Mature Lodgepole Pine (Pinus Contorta) and Adjacent Stands of Old Growth Mixed Conifer in Yellowstone National Park, Wyoming USA

Forest development patterns following disturbance are known to influence the physical and chemical attributes of soils at different points in time. Changes in soil resources are thought to have a corresponding effect on ectomycorrhizal (ECM) community structure. We used molecular methods to compare below-ground ECM species richness, composition, and abundance between adjacent stands of homogenous lodgepole pine and old growth mixed conifer in Yellowstone National Park (YNP). In each stand-type we collected soil cores to both identify mycorrhizae and assess soil chemistry. Although no statistical difference was observed in the mean number of ECM root tips per core between stand types, the total number of species identified (85 versus 35) and the mean number of species per core (8.8 +/- 0.6 versus 2.5 +/- 0.3) were significantly higher in lodgepole pine. Differences between the actual and estimated species richness levels indicated that these forest types support a high number of ECM species and that undersampling was severe. Species compositions were widely disparate between stands where only four species were shared out of a total of 116. Soil analysis also revealed that mixed conifer was significantly lower in pH, but higher in organic matter, potassium, phosphorus, and ammonium when compared to lodgepole pine stands. Species richness per core was correlated with these chemical data, however, analysis of covariance indicated that stand type was the only statistically significant factor in the observed difference in species richness. Our data suggest that ECM fungal richness increases as homogenous lodgepole pine stands grow and mature, but declines after Engelmann spruce and subalpine fir colonize. Despite difficulties linking species composition with soil chemistry, there are a variety of physical and chemical factors that could be influencing ECM community structure. Future field experiments are necessary to test some of the mechanisms potentially operating within this system.

Douglas, Robert B.

Effects of extracellular matrix proteins on macrophage differentiation, growth, and function: comparison of liquid and agar culture systems

Both spaceflight and skeletal unloading suppress the haematopoietic differentiation of macrophages (Sonnenfeld et al., Aviat. Space Environ. Med., 61:648-653, 1990; Armstrong et al., J. Appl. Physiol., 75:2734-2739, 1993). The mechanism behind this reduction in haematopoiesis has yet to be elucidated. However, changes in bone marrow extracellular matrix (ECM) may be involved. To further understand the role of ECM products in macrophage differentiation, we have performed experiments evaluating the effects of fibronectin, laminin, collagen type I, and collagen type IV on macrophage development and function. Bone marrow-derived macrophages cultured on four different ECM substrates in liquid culture medium showed less growth than those cultured on plastic. Significant morphological differences were seen on each of the substrates used. Phenotypically and functionally, as measured by class II major histocompatibility molecule (MHCII) expression, MAC-2 expression, and the secretion of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), these macrophages were similar. In contrast, bone marrow-derived macrophages cultured in suspension, using agar, showed no difference in growth when exposed to ECM proteins. However, IL-6 and TNF-alpha secretion was affected by fibronectin, laminin, collagen type I, and collagen type IV in a concentration-dependent manner. We conclude that the ECM products fibronectin, laminin, collagen type I, and collagen type IV have profound effects on macrophage development and function. Additionally, we suggest that an ECM-supplemented agar culture system provides an environment more analogous to in vivo bone marrow than does a traditional liquid culture system.

Non-NASA Center

Integrin binding and mechanical tension induce movement of mRNA and ribosomes to focal adhesions

The extracellular matrix (ECM) activates signalling pathways that control cell behaviour by binding to cell-surface integrin receptors and inducing the formation of focal adhesion complexes (FACs). In addition to clustered integrins, FACs contain proteins that mechanically couple the integrins to the cytoskeleton and to immobilized signal-transducing molecules. Cell adhesion to the ECM also induces a rapid increase in the translation of preexisting messenger RNAs. Gene expression can be controlled locally by targeting mRNAs to specialized cytoskeletal domains. Here we investigate whether cell binding to the ECM promotes formation of a cytoskeletal microcompartment specialized for translational control at the site of integrin binding. High-resolution in situ hybridization revealed that mRNA and ribosomes rapidly and specifically localized to FACs that form when cells bind to ECM-coated microbeads. Relocation of these protein synthesis components to the FAC depended on the ability of integrins to mechanically couple the ECM to the contractile cytoskeleton and on associated tension-moulding of the actin lattice. Our results suggest a new type of gene regulation by integrins and by mechanical stress which may involve translation of mRNAs into proteins near the sites of signal reception.

Non-NASA Center

Extracellular matrix controls tubulin monomer levels in hepatocytes by regulating protein turnover

Cells have evolved an autoregulatory mechanism to dampen variations in the concentration of tubulin monomer that is available to polymerize into microtubules (MTs), a process that is known as tubulin autoregulation. However, thermodynamic analysis of MT polymerization predicts that the concentration of free tubulin monomer must vary if MTs are to remain stable under different mechanical loads that result from changes in cell adhesion to the extracellular matrix (ECM). To determine how these seemingly contradictory regulatory mechanisms coexist in cells, we measured changes in the masses of tubulin monomer and polymer that resulted from altering cell-ECM contacts. Primary rat hepatocytes were cultured in chemically defined medium on bacteriological petri dishes that were precoated with different densities of laminin (LM). Increasing the LM density from low to high (1-1000 ng/cm2), promoted cell spreading (average projected cell area increased from 1200 to 6000 microns2) and resulted in formation of a greatly extended MT network. Nevertheless, the steady-state mass of tubulin polymer was similar at 48 h, regardless of cell shape or ECM density. In contrast, round hepatocytes on low LM contained a threefold higher mass of tubulin monomer when compared with spread cells on high LM. Furthermore, similar results were obtained whether LM, fibronectin, or type I collagen were used for cell attachment. Tubulin autoregulation appeared to function normally in these cells because tubulin mRNA levels and protein synthetic rates were greatly depressed in round cells that contained the highest level of free tubulin monomer. However, the rate of tubulin protein degradation slowed, causing the tubulin half-life to increase from approximately 24 to 55 h as the LM density was lowered from high to low and cell rounding was promoted. These results indicate that the set-point for the tubulin monomer mass in hepatocytes can be regulated by altering the density of ECM contacts and changing cell shape. This finding is consistent with a mechanism of MT regulation in which the ECM stabilizes MTs by both accepting transfer of mechanical loads and altering tubulin degradation in cells that continue to autoregulate tubulin synthesis.

Non-NASA Center

Control of cytoskeletal mechanics by extracellular matrix, cell shape, and mechanical tension

We have investigated how extracellular matrix (ECM) alters the mechanical properties of the cytoskeleton (CSK). Mechanical stresses were applied to integrin receptors on the apical surfaces of adherent endothelial cells using RGD-coated ferromagnetic microbeads (5.5-microns diameter) in conjunction with a magnetic twisting device. Increasing the number of basal cell-ECM contacts by raising the fibronectin (FN) coating density from 10 to 500 ng/cm2 promoted cell spreading by fivefold and increased CSK stiffness, apparent viscosity, and permanent deformation all by more than twofold, as measured in response to maximal stress (40 dyne/cm2). When the applied stress was increased from 7 to 40 dyne/cm2, the stiffness and apparent viscosity of the CSK increased in parallel, although cell shape, ECM contacts, nor permanent deformation was altered. Application of the same stresses over a lower number ECM contacts using smaller beads (1.4-microns diameter) resulted in decreased CSK stiffness and apparent viscosity, confirming that this technique probes into the depth of the CSK and not just the cortical membrane. When magnetic measurements were carried out using cells whose membranes were disrupted and ATP stores depleted using saponin, CSK stiffness and apparent viscosity were found to rise by approximately 20%, whereas permanent deformation decreased by more than half. Addition of ATP (250 microM) under conditions that promote CSK tension generation in membrane-permeabilized cells resulted in decreases in CSK stiffness and apparent viscosity that could be detected within 2 min after ATP addition, before any measurable change in cell size.(ABSTRACT TRUNCATED AT 250 WORDS).

Non-NASA Center

Extracellular matrix as a solid-state regulator in angiogenesis: identification of new targets for anti-cancer therapy

Angiogenesis, the growth of blood capillaries, is regulated by soluble growth factors and insoluble extracellular matrix (ECM) molecules. Soluble angiogenic mitogens act over large distances to initiate capillary growth whereas changes in ECM govern whether individual cells will grow, differentiate, or involute in response to these stimuli in the local tissue microenvironment. Analysis of this local control mechanism has revealed that ECM molecules switch capillary endothelial cells between differentiation and growth by both binding specific transmembrane integrin receptors and physically resisting cell-generated mechanical loads that are applied to these receptors. Control of capillary endothelial cell form and function therefore may be exerted by altering the mechanical properties of the ECM as well as its chemical composition. Understanding of this mechanochemical control mechanism has led to the development of new angiogenesis inhibitors that may be useful for the treatment of cancer.

Review

Extracellular matrix and cell shape: potential control points for inhibition of angiogenesis

Capillary endothelial (CE) cells require two extracellular signals in order to switch from quiescence to growth and back to differentiation during angiogenesis: soluble angiogenic factors and insoluble extracellular matrix (ECM) molecules. Soluble endothelial mitogens, such as basic fibroblast growth factor (FGF), act over large distances to trigger capillary growth, whereas ECM molecules act locally to modulate cell responsiveness to these soluble cues. Recent studies reveal that ECM molecules regulate CE cell growth and differentiation by modulating cell shape and by activating intracellular chemical signaling pathways inside the cell. Recognition of the importance of ECM and cell shape during capillary morphogenesis has led to the identification of a series of new angiogenesis inhibitors. Elucidation of the molecular mechanism of capillary regulation may result in development of even more potent angiogenesis modulators in the future.

Non-NASA Center

Localization of extracellular matrix components in developing mouse salivary glands by confocal microscopy

The importance of the extracellular matrix (ECM) in epithelial-mesenchymal interactions in developing organisms is well established. Proteoglycans and interstitial collagens are required for the growth, morphogenesis, and differentiation of epithelial organs and the distribution of these molecules has been described. However, much less is known about other ECM macromolecules in developing epithelial organs. We used confocal microscopy to examine the distribution of laminin, heparan sulfate (BM-1) proteoglycan, fibronectin, and collagen types I, IV, and V, in mouse embryonic salivary glands. Organ rudiments were isolated from gestational day 13 mouse embryos and cultured for 24, 48, or 72 hours. Whole mounts were stained by indirect immunofluorescence and then examined using a Zeiss Laser Scan Microscope. We found that each ECM component examined had a distinct distribution and that the distribution of some molecules varied with culture time. Laminin was mainly restricted to the basement membrane. BM-1 proteoglycan was concentrated in the basement membrane and also formed a fine network throughout the mesenchyme. Type IV collagen was mainly located in the basement membrane of the epithelium, but it was also present throughout the mesenchyme. Type V collagen was distributed throughout the mesenchyme at 24 hours, but at 48 hours was principally located in the basement membrane. Type I collagen was distributed throughout the mesenchyme at all culture times, and accumulated in the clefts and particularly at the epithelial-mesenchymal interface as time in culture increased. Fibronectin was observed throughout the mesenchyme at all times.

Hardman, P.

Electron cyclotron maser emission at oblique angles

Possible causes of observations of electron cyclotron maser emissions (ECMEs) at oblique angle (about 60 deg) to the magnetic field are investigated. The paper discusses general concepts of ECME in terms of resonant ellipses and considers electron distributions required to produce ECME at oblique angles, as well as the ways in which the most favorable of these distributions may be produced. A mechanism is proposed that might produce an appropriate 'spiraling beam' distribution, with a peak in velocity space at speed v - v(0) and pitch angle alpha = alpha(0) not equal to 0.

Melrose, D. B.

Experiment K-7-29: Connective Tissue Studies: Changes in Muscle Serine Proteases, Serpins and Matrix Molecules - Part 2

In zero or micro-gravity, type 1 muscle fibers atrophy and lose predominance, especially in slow-twitch muscles. No increase in mononuclear cells has been observed, just as in simple denervation, where both types 1 and 2 fibers atrophy, again without infiltration of cells, but with clear satellite cell proliferation. However, extracellular matrix (ECM) degradation takes place after denervation and if re-innervation is encouraged, functional recovery to near control levels may be achieved. No information is available concerning the ECM milieu, the activation of serine proteases, their efficacy in degrading ECM components and the production of locally-derived natural protease inhibitors (serpins) in effecting surface proteolytic control. In addition, no studies are available concerning the activation of these enzymes in micro- or zero gravity or their response to muscle injury on the ground and what alterations, if any, occur in space. These studies were the basis for the experiments in Cosmos 2044.

Festoff, B. W.

Directional control of lamellipodia extension by constraining cell shape and orienting cell tractional forces

Directed cell migration is critical for tissue morphogenesis and wound healing, but the mechanism of directional control is poorly understood. Here we show that the direction in which cells extend their leading edge can be controlled by constraining cell shape using micrometer-sized extracellular matrix (ECM) islands. When cultured on square ECM islands in the presence of motility factors, cells preferentially extended lamellipodia, filopodia, and microspikes from their corners. Square cells reoriented their stress fibers and focal adhesions so that tractional forces were concentrated in these corner regions. When cell tension was dissipated, lamellipodia extension ceased. Mechanical interactions between cells and ECM that modulate cytoskeletal tension may therefore play a key role in the control of directional cell motility.

NASA Discipline Cell Biology

Mechanotransduction: all signals point to cytoskeleton, matrix, and integrins

Mechanical stresses modulate cell function by either activating or tuning signal transduction pathways. Mechanotransduction, the process by which cells convert mechanical stimuli into a chemical response, occurs both in cells specialized for sensing mechanical cues and in parenchymal cells whose primary function is not mechanosensory. However, common among the various responses to mechanical stress is the importance of direct or indirect connections between the internal cytoskeleton, the extracellular matrix (ECM), and traditional signal transducing molecules. In many instances, these elements converge at focal adhesions, sites of structural attachment between the cytoskeleton and ECM that are anchored by cell surface integrin receptors. Alenghat and Ingber discuss the accumulating evidence for the central role of cytoskeleton, ECM, and integrin-anchored focal adhesions in several mechanotransduction pathways.

Review, Tutorial

Integrin-extracellular matrix interactions in connective tissue remodeling and osteoblast differentiation

The differentiaton of bone cells is a complex multistep process. Bone is somewhat unusual in that it is very actively and continually remodeled in the adult and that maintenance of its mass in the mature organism is exquisitely sensitive to mechanical as well as chemical signals. Bone is also unique because it consists of a very large amount of extracellular matrix (ECM) that is mineralized. The integrin family of ECM receptors has been shown to play an important role in tissue morphogenesis in several systems. Our studies on the regulation of matrix remodeling enzymes by integrins in rabbit synovial fibroblasts show that two b1 integrin fibronectin (FN) receptor complexes (alpha 5 beta 1 and alpha 4 beta 1) cooperate in detecting subtle changes in the composition of the ECM. As a result of signal transduction by these integrins, the levels of mRNA and protein for several members of the metalloproteinase family are regulated in these cells. We have also used antibody and RGD peptide perturbation studies to determine the significance of cell/ECM interactions to normal osteogenesis. We found that interactions between the cell binding domain of FN and integrins are required for both normal morphogenesis and gene expression in cultured osteoblasts that differentiate to form bone-like tissue in culture. These data lead us to propose that beta 1 integrins play an important role in osteoblast differentiation as well as in bone remodeling.

NASA Discipline Number 26-10

Growth and morphogenesis of embryonic mouse organs on non-coated and extracellular matrix-coated Biopore membrane

Embryonic mouse salivary glands, pancreata, and kidneys were isolated from embryos of appropriate gestational age by microdissection, and were cultured on Biopore membrane either non-coated or coated with type I collagen or Matrigel. As expected, use of Biopore membrane allowed high quality photomicroscopy of the living organs. In all organs extensive mesenchymal spreading was observed in the presence of type I collagen or Matrigel. However, differences were noted in the effects of extracellular matrix (ECM) coatings on epithelial growth and morphogenesis: salivary glands were minimally affected, pancreas morphogenesis was adversely affected, and kidney growth and branching apparently was enhanced. It is suggested that these differences in behaviour reflect differences in the strength of interactions between the mesenchymal cells and their surrounding endogenous matrix, compared to the exogenous ECM macromolecules. This method will be useful for culture of these and other embryonic organs. In particular, culture of kidney rudiments on ECM-coated Biopore offers a great improvement over previously used methods which do not allow morphogenesis to be followed in vitro.

NASA Discipline Number 93-10

Semicarbazide-sensitive amine oxidase and extracellular matrix deposition by smooth-muscle cells

We have recently reported in vivo disruption of collagen and elastin architecture within blood vessel walls resulting from the selective inhibition of the enzyme semicarbazide-sensitive amine oxidase (SSAO). This study further investigates the effects of SSAO inhibition on extracellular matrix deposition by smooth-muscle cells (SMCs) cultured from neonatal rat hearts. SMCs were characterized, SSAO activity was measured, and soluble and insoluble collagen and elastin in the extracellular matrix (ECM) were quantified. Cultured neonatal rat heart SMC exhibited a monotypic synthetic phenotype that likely represents a myofibroblast. Detectable levels of SSAO activity present throughout 30-d culture peaked at 7-14 d, coinciding with the production of ECM. The addition of enzyme inhibitors and alternate SSAO substrates (benzylamine) produced varied and, in some cases, marked changes in SSAO activity as well as in the composition of mature and soluble matrix components. Similar to our previous in vivo findings, in vitro SSAO inhibition produced aberrations in collagen and elastin deposition by heart SMC. Because changes in SSAO activity are associated with cardiovascular pathologic states, this enzyme may play a protective or modulating role by regulating ECM production during pathologic insult.

Allylamine/administration & dosage/analogs & deriv