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At least 55 records · Page 3
All sourcesNASA NTRSDOE OSTI
NASA NTRS1997

Activation of G proteins mediates flow-induced prostaglandin E2 production in osteoblasts

Interstitial fluid flow may play a role in load-induced bone remodeling. Previously, we have shown that fluid flow stimulates osteoblast production of cAMP inositol trisphosphate (IP3), and PGE2. Flow-induced increases in cAMP and IP3 were shown to be a result of PG production. Thus, PGE2 production appears to be an important component in fluid flow induced signal transduction. In the present study, we investigated the mechanism of flow-induced PGE2 synthesis. Flow-induced a 20-fold increase in PGE2 production in osteoblasts. Increases were also observed with ALF4-(10mM) (98-fold), an activator of guanidine nucleotide-binding proteins (G proteins), and calcium ionophore A23187 (2 microM) (100-fold) in stationary cells. We then investigated whether flow stimulation is mediated by G proteins and increases in intracellular calcium. Flow-induced PGE2 production was inhibited by the G protein inhibitors GDP beta S (100 microM) and pertussis toxin (1 microgram/ml) by 83% and 72%, respectively. Chelation of extracellular calcium by EGTA (2 mM) and intracellular calcium by quin-2/AM (30 microM) blocked flow stimulation by 87% and 67%, respectively. These results suggest that G proteins and calcium play an important role in mediating mechanochemical signal transduction in osteoblasts.

NASA Discipline Cell Biology↗
NASA NTRS1996

Redox-mediated activation of latent transforming growth factor-beta 1

Transforming growth factor beta 1 (TGF beta) is a multifunctional cytokine that orchestrates response to injury via ubiquitous cell surface receptors. The biological activity of TGF beta is restrained by its secretion as a latent complex (LTGF beta) such that activation determines the extent of TGF beta activity during physiological and pathological events. TGF beta action has been implicated in a variety of reactive oxygen-mediated tissue processes, particularly inflammation, and in pathologies such as reperfusion injury, rheumatoid arthritis, and atherosclerosis. It was recently shown to be rapidly activated after in vivo radiation exposure, which also generates reactive oxygen species (ROS). In the present studies, the potential for redox-mediated LTGF beta activation was investigated using a cell-free system in which ROS were generated in solution by ionizing radiation or metal ion-catalyzed ascorbate reaction. Irradiation (100 Gray) of recombinant human LTGF beta in solution induced 26% activation compared with that elicited by standard thermal activation. Metal-catalyzed ascorbate oxidation elicited extremely efficient recombinant LTGF beta activation that matched or exceeded thermal activation. The efficiency of ascorbate activation depended on ascorbate concentrations and the presence of transition metal ions. We postulate that oxidation of specific amino acids in the latency-conferring peptide leads to a conformation change in the latent complex that allows release of TGF beta. Oxidative activation offers a novel route for the involvement of TGF beta in tissue processes in which ROS are implicated and endows LTGF beta with the ability to act as a sensor of oxidative stress and, by releasing TGF beta, to function as a signal for orchestrating the response of multiple cell types. LTGF beta redox sensitivity is presumably directed toward recovery of homeostasis; however, oxidation may also be a mechanism of LTGF beta activation that can be deleterious during disease mechanisms involving chronic ROS production.

Non-NASA Center↗
NASA NTRS2013

Atomic Force Microscope Mediated Chromatography

The atomic force microscope (AFM) is used to inject a sample, provide shear-driven liquid flow over a functionalized substrate, and detect separated components. This is demonstrated using lipophilic dyes and normal phase chromatography. A significant reduction in both size and separation time scales is achieved with a 25-micron-length column scale, and one-second separation times. The approach has general applications to trace chemical and microfluidic analysis. The AFM is now a common tool for ultra-microscopy and nanotechnology. It has also been demonstrated to provide a number of microfluidic functions necessary for miniaturized chromatography. These include injection of sub-femtoliter samples, fluidic switching, and sheardriven pumping. The AFM probe tip can be used to selectively remove surface layers for subsequent microchemical analysis using infrared and tip-enhanced Raman spectroscopy. With its ability to image individual atoms, the AFM is a remarkably sensitive detector that can be used to detect separated components. These diverse functional components of microfluidic manipulation have been combined in this work to demonstrate AFM mediated chromatography. AFM mediated chromatography uses channel-less, shear-driven pumping. This is demonstrated with a thin, aluminum oxide substrate and a non-polar solvent system to separate a mixture of lipophilic dyes. In conventional chromatographic terms, this is analogous to thin-layer chromatography using normal phase alumina substrate with sheardriven pumping provided by the AFM tip-cantilever mechanism. The AFM detection of separated components is accomplished by exploiting the variation in the localized friction of the separated components. The AFM tip-cantilever provides the mechanism for producing shear-induced flows and rapid pumping. Shear-driven chromatography (SDC) is a relatively new concept that overcomes the speed and miniaturization limitations of conventional liquid chromatography. SDC is based on a sliding plate system, consisting of two flat surfaces, one of which has a recessed channel. A fluid flow is produced by axially sliding one plate past another, where the fluid has mechanical shear forces imposed at each point along the channel length. The shear-induced flow rates are very reproducible, and do not have pressure or voltage gradient limitations. SDC opens up a new range of enhanced separation kinetics by permitting the sample confinement with submicron dimensions. Small, highly confined liquid is advantageous for chromatographic separation because the separation rate is known to scale according to the square of the confined sample diameter. In addition, because shear-driven flows are not limited by fluid velocity, shear-driven liquid chromatography may provide up to 100,000 plate efficiency.

Anderson, Mark S.↗
NASA NTRSDate not supplied

A Dominant Arabidopsis Thaliana ACTIN7 Mutant for Studies of Cytoskeletal-Mediated Root Gravity and Spaceflight Stress Responses

A forward genetic screen for Arabidopsis thaliana mutants that exhibited differential sensitivity to the actin-disrupting compound, latrunculin B (LatB), was conducted to uncover new players involved in actin-mediated root gravity responses. This to the isolation of a mutant that exhibited robust primary root growth at 100 nM LatB, which is a concentration that severely inhibits wild-type root elongation. Phenotypic analysis revealed that hypocotyl elongation in the dark and root hair tip growth in the mutant could tolerate LatB concentrations that impaired these processes in wild type. A cross between the mutant and wild type resulted in progeny resistance to LatB, which indicated that the mutant is dominant (hereafter referred to as LBR1 for LatB Resistant1). Filamentous-actin (F-actin) organization in LBR1 primary roots remained intact at 100 nM LatB, while that of wild type exhibited fragmented F-actin. Next generation sequencing revealed that LBR1 had a single nucleotide polymorphism (SNP) in the AT5G09810 gene that changed cytosine to a thiamine at the first exon. AT5G09810 encodes ACTIN7 (ACT7), which is one of three vegetative actin isoforms in A. thaliana. The SNP in the ACT7 gene led to a change in a single amino acid from proline at position 32 to a serine. Transgenic complementation of LBR1 plants with wild-type ACT7 under the control of the ACT7 promoter (pACT7:ACT7) and wild-type plants with LBR1, which contained the proline to serine mutation, also under the ACT7 promoter (pACT7:LBR1) confirmed that LBR1 is ACT7. The pACT7:LBR1 construct was also able to confer LatB resistance to the act7-5 and act2-3 vegetative ACT mutants. Moreover, LBR1 exhibited partial tolerance to salt and low phosphate, and enhanced root skewing on a clinostat, suggesting that site-directed engineering of vegetative ACT presents a strategy for generating stress-tolerant plants for spaceflight applications and studies of actin-mediated gravity responses.

Plant Space Biology↗
NASA NTRSDate not supplied

A Dominant Arabidopsis Thaliana ACTIN7 Mutant for Studies of Cytoskeletal-Mediated Root Gravity and Spaceflight Stress Responses

A forward genetic screen for Arabidopsis thaliana mutants that exhibited differential sensitivity to the actin-disrupting compound, latrunculin B (LatB), was conducted to uncover new players involved in actin-mediated root gravity responses. This led to the isolation of a mutant that exhibited robust primary root growth at 100 nM LatB, which is a concentration that severely inhibits wild-type root elongation. Phenotypic analysis revealed that hypocotyl elongation in the dark and root hair tip growth in the mutant could tolerate LatB concentrations that impaired these processes in wild type. A cross between the mutant and wild type resulted in progeny resistance to LatB, which indicated that the mutant is dominant (hereafter referred to as LBR1 for LatB Resistant1). Filamentous-actin (F-actin) organization in LBR1 primary roots remained intact at 100 nM LatB, while that of wild type exhibited fragmented F-actin. Next generation sequencing revealed that LBR1 had a single nucleotide polymorphism (SNP) in the AT5G09810 gene that changed cytosine to a thiamine at the first exon. AT5G09810 encodes ACTIN7 (ACT7), which is one of three vegetative actin isoforms in A. thaliana. The SNP in the ACT7 gene led to a change in a single amino acid from proline at position 32 to a serine. Transgenic complementation of LBR1 plants with wild-type ACT7 under the control of the ACT7 promoter (pACT7:ACT7) and wild-type plants with LBR1, which contained the proline to serine mutation, also under the ACT7 promoter (pACT7:LBR1) confirmed that LBR1 is ACT7. The pACT7:LBR1 construct was also able to confer LatB resistance to the act7-5 and act2-3 vegetative ACT mutants. Moreover, LBR1 exhibited partial tolerance to salt and low phosphate, and enhanced root skewing on a clinostat, suggesting that site-directed engineering of vegetative ACT presents a strategy for generating stress-tolerant plants for spaceflight applications and studies of actin-mediated gravity responses.

Plant Space Biology↗
NASA NTRSDate not supplied

A Pair of Trans-Golgi Network/Early Endosome-Localized Proteins Facilitate Cytoskeletal-Mediated Root Skewing in Arabidopsis Thaliana

Roots treated with the actin-disrupting compound Latrunculin B (LatB) show stronger gravitropic responses on Earth and dampened straightening responses on a 2-D clinostat. In related experiments using the Biological Research in Canisters (BRIC) hardware, it was found that knockouts to vegetative actin isoforms in Arabidopsis thaliana had more robust root skewing in microgravity and waved more strongly than wild type on hard agar surfaces. These results indicate that the actin cytoskeleton mediates directional root growth in space and on the ground. To gain new insights into the role of actin in directional root growth, we identified mutants that showed differential growth responses to low doses of LatB. In-depth studies of one mutant led to the identification of a trans-Golgi Network (TGN)/Early Endosome (EE)- localized protein that associates with actin. This protein called hypersensitive to LatB 1 (HLB1) colocalized with the ADP-ribosylation-factor guanine nucleotide exchange factor, MIN7/BEN1 (HOPM INTERACTOR7/BREFELDIN A-VISUALIZED ENDOCYTIC TRAFFICKING DEFECTIVE1), at the TGN/EE. HLB1 and MIN7/BEN1 were found to regulate exocytosis and endocytosis, respectively, suggesting that both proteins are involved in actin-mediated membrane traffic. Microtubules, another component of the cytoskeleton, is known to be involved in root skewing. Both hlb1 and min7/ben1 mutants exhibited dampened root skewing on the microtubule stabilizing compound taxol. Taken together, our results support the conclusion that cytoskeletal-membrane interactions contribute to directional root skewing in A. thaliana and is facilitated in part by the TGN/EE-localized HLB1 and MIN7/BEN1 proteins.

Plant Space Biology↗
NASA NTRS1976

Compensatory adrenal growth - A neurally mediated reflex

The responses of young rats to left adrenalectomy or left adrenal manipulation were compared to surgical sham adrenalectomy in which adrenals were observed but not touched. At 12 h right adrenal wet weight, dry weight, DNA, RNA, and protein content were increased (P less than 0.05) after the first two operations. Left adrenal manipulation resulted in increased right adrenal weight at 12 h but no change in left adrenal weight. Sequential manipulation of the left adrenal at time 0 and the right adrenal at 12 h resulted in an enlarged right adrenal at 12 h (P less than 0.01), and an enlarged left adrenal at 24 h (P less than 0.05), showing that the manipulated gland was capable of response. Bilateral adrenal manipulation of the adrenal glands resulted in bilateral enlargement of 12 h (P less than 0.01). Taken together with previous results, these findings strongly suggest that compensatory adrenal growth is a neurally mediated reflex.

Dallman, M. F.↗
NASA NTRS1977

Cortisol-mediated synchronization of circadian rhythm in urinary potassium excretion

Conscious chair-acclimatized squirrel monkeys (Saimiri sciureus) studied with lights on (600 lx) from 0800 to 2000 hr daily (LD 12:12) display a prominent circadian rhythm in renal potassium excretion. The characteristics of this rhythm were reproduced in adrenalectomized monkeys by infusing 5 mg cortisol and 0.001 mg aldosterone, or 5 mg cortisol alone, between 0800 and 0900 kr daily. When the timing of cortisol administration (with or without aldosterone) was phase-delayed by 8 hr, the urinary potassium rhythm resynchronized by 80% of the cortisol phase shift, but only after a transient response lasting 3-4 days. With the same daily dose of adrenal steroids given as a continuous infusion throughout each 24 hr, urinary potassium excretion showed free-running oscillations no longer synchronized to the light-dark cycle. These results indicate that the circadian rhythm of plasma cortisol concentration acts as an internal mediator in the circadian timing system, synchronizing a potentially autonomous oscillation in renal potassium excretion to environmental time cues and to other circadian rhythms within the animal.

Moore-Ede, M. C.↗
NASA NTRS1982

Evaluation of the sensitivity and intrusion of workload estimation techniques in piloting tasks emphasizing mediational activity

In this experiment, pilots flew an instrumented moving-base simulator. Mediational loading was elicited by having them solve a variety of navigational problems. The problems were sorted into low, medium, and high load conditions based on the number and complexity of arithmetic and geometric operations required to solve them. Workload estimation techniques based on opinion, spare mental capacity, primary task performance, and physiological measures were obtained and compared. This paper describes: (1) the ability of the techniques to discriminate statistically between the three levels of loading conditions, and (2) changes in primary task performance caused by introduction of the workload technique procedures and equipment.

Rahimi, M.↗
NASA NTRS1984

Rheoceptive mediators of graviperception in a water flea: Morphological implications of antennal-socket setae in daphnia magna

Aquatic microcrustaceans of the genus Daphnia are known to orient to light during the day. At night, in the absence of visual cues, daphnids were suspected of maintaining equilibrium by monitoring the direction of gravity through their swimming antennae. Recent investigations using simulated, weightlessness conditions coupled with absence of illumination revealed hair like structures or setae on the basal, articulating socket of the antennae that, when surgically removed, resulted in disorientation. Given the simulated weightlessness or neutrally buoyant condition that eliminated sinking of the normally negatively buoyant Daphnia, it was proposed that the antennal socket setae function as rheoceptors stimulated by the upward rush of water currents during gravity induced, sinking phase of daphnid swimming movements. This rheoceptively mediated, gravity perception hypothesis is further supported by morphological investigations. Scanning electron micrographs indicate that antennal socket setae are anatomically similar to proprioceptors used by higher crustaceans to monitor gravitational direction.

Meyers, D. G.↗
NASA NTRS1983

Monte Carlo simulation of steady state shock structure including cosmic ray mediation and particle escape

Both hydrodynamic calculations (Drury and Volk, 1981, and Axford et al., 1982) and kinetic simulations imply the existence of thermal subshocks in high-Mach-number cosmic-ray-mediated shocks. The injection efficiency of particles from the thermal background into the diffusive shock-acceleration process is determined in part by the sharpness and compression ratio of these subshocks. Results are reported for a Monte Carlo simulation that includes both the back reaction of accelerated particles on the inflowing plasma, producing a smoothing of the shock transition, and the free escape of particles allowing arbitrarily large overall compression ratios in high-Mach-number steady-state shocks. Energy spectra and estimates of the proportion of thermal ions accelerated to high energy are obtained.

Ellison, D. C.↗
NASA NTRS1985

Hyper-G stress-induced hyperglycemia in rats mediated by glucoregulatory hormones

The present investigation is concerned with possible relations of the hyperglycemic response of rats exposed to hyper-G stress to (1) alterations in blood levels of the glucoregulatory hormones and gluconeogenic substrates, and (2) changes in insulin response on muscle glucose uptake. Male Sprague-Dawley rats weighing 250-300 g were used in the study. The results of the experiments indicate that the initial rapid rise in blood glucose of rats exposed to hyper-G stress is mediated by increases in circulating catecholamines and glucagon, both potent stimulators of hepatic gluconeogenesis. Lactate, derived from epinephrine stimulation of muscle glycogenolysis, appears to be a major precursor for the initial rise in blood glucose. The inhibition of the insulin-stimulated glucose uptake by muscle tissues may be a factor in the observed sustained hyperglycemia.

Daligcon, B. C.↗
NASA NTRS1985

Bone loss during simulated weightlessness - Is it glucocorticoid mediated?

Elevating the hindquarters of a rat by the tail unweights the hind limbs but maintains normal weight-bearing by the forelimbs. This maneuver leads to a decrease in bone mass and calcium content in the unweighted bones (e.g., tibia and L1 vertebra), but not in the normally weighted bones (e.g., humerus and mandible). Potentially, the stress of the maneuver, mediated by increased glucocorticoid production and secretion, could explain the decreased bone formation, rather than the skeletal unweighting per se. To test this possibility, the effects of adrenalectomy on the response of bone to the unweighting of the hind limbs of normal rats were evaluated.

Bikle, D. D.↗