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At least 343 records · Page 19

Control of intracellular pH and growth by fibronectin in capillary endothelial cells

The aim of this work was to analyze the mechanism by which fibronectin (FN) regulates capillary endothelial cell proliferation. Endothelial cell growth can be controlled in chemically-defined medium by varying the density of FN coated on the substratum (Ingber, D. E., and J. Folkman. J. Cell Biol. 1989. 109:317-330). In this system, DNA synthetic rates are stimulated by FN in direct proportion to its effect on cell extension (projected cell areas) both in the presence and absence of saturating amounts of basic FGF. To investigate direct growth signaling by FN, we carried out microfluorometric measurements of intracellular pH (pHi), a cytoplasmic signal that is commonly influenced by soluble mitogens. pHi increased 0.18 pH units as FN coating densities were raised and cells progressed from round to spread. Intracellular alkalinization induced by attachment to FN was rapid and followed the time course of cell spreading. When measured in the presence and absence of FGF, the effects of FN and FGF on pHi were found to be independent and additive. Furthermore, DNA synthesis correlated with pHi for all combinations of FGF and FN. Ethylisopropylamiloride, a specific inhibitor of the plasma membrane Na+/H+ antiporter, completely suppressed the effects of FN on both pHi and DNA synthesis. However, cytoplasmic pH per se did not appear to be a critical determinant of growth since DNA synthesis was not significantly inhibited when pHi was lowered over the physiological range by varying the pH of the medium. We conclude that FN and FGF exert their growth-modulating effects in part through activation of the Na+/H+ exchanger, although they appear to trigger this system via separate pathways.

NASA Discipline Cell Biology↗

Human mononuclear cell function after 4 degrees C storage during 1-G and microgravity conditions of spaceflight

Future space missions of long duration may require that autologous leukocytes be stored in flight for infusion to restore normal immune competence in crewmembers. Peripheral blood mononuclear cells (PBMNCs), as leukocyte concentrates in autologous plasma and 2% dextrose, were stored in the microgravity conditions provided by the U.S. Space Shuttle Columbia (Mission 61-C). Activity of PBMNC after space flight was compared with that from a series of preflight ground control experiments, which demonstrated in culture a progressive daily loss in mitogen-stimulated protein synthesis at 24 h and thymidine uptake at 72 h after storage for 7 d at 4 degrees C. Post-storage viabilities were at least 90% as determined by trypan dye exclusion. A progressive reduction in the percentage of PBMNC expressing cell-surface phenotype markers, which was similar for monocytes, B cells, and T-cell subsets, also occurred after storage. The ability of PBMNC, stored for 8 d in Columbia's middeck, to become activated and proliferate in vitro was similar to that of cells that remained in identical flight lockers on the ground as 1-G controls, thus indicating that PBMNCs were not adversely affected by storage under microgravity conditions.

NASA Discipline Number 00-00↗

Myoneural necrosis following high-frequency electrical stimulation of the cast-immobilized rabbit hindlimb

The morphological and physiological effects of 4 weeks of high-frequency electrical stimulation (1 h/day, 5 days/week) on cast-immobilized rabbit hindlimbs were investigated in the tibialis anterior muscle and peroneal nerve. In 2 out of 6 animals, high-frequency stimulation with immobilization caused muscle fiber death, internalization of muscle fiber nuclei, connective tissue proliferation, inflammatory response, altered fiber size distribution and variable staining intensities. The fast-twitch fibers were predominantly affected. Two of six peripheral nerves subjected to immobilization and stimulation showed severe damage. Tetanic forces were significantly reduced in the affected muscles. Therefore, the immobilization and high-frequency stimulation may be detrimental to myoneural structure and function and, thus, this combination of therapies should be applied conservatively.

Non-NASA Center↗

Nuclear size as a cell-kinetic marker for osteoblast differentiation

A nuclear morphometric assay for preosteoblasts is introduced as a cell-kinetic technique, applicable to routine histological preparations of mineralized tissue. Because this method is a morphological marker for osteoblast precursor cell differentiation, it provides a new dimension for determining the mechanism of osteoblast histogenesis. Osteoblast precursors of the periodontal ligament are a mixed population of progenitors, kinetically separable into two distinct groups according to nuclear size. Preosteoblasts, the immediate proliferating precursors of osteoblasts, have large nuclei (greater than 170 micrometers3) and are derived from relatively undifferentiated fibroblastlike cells, which have smaller nuclei (less than 80 micrometers3). Increase in nuclear volume, during G1 phase of the cell cycle, is apparently a morphological manifestation of change in genomic expression. This key event in preosteoblast differentiation is related to mechanical stress/strain and may be an important rate-limiting step in osteoblast histogenesis.

Non-NASA Center↗

Morphologic differentiation of colon carcinoma cell lines HT-29 and HT-29KM in rotating-wall vessels

A new low shear stress microcarrier culture system has been developed at NASA's Johnson Space Center that permits three-dimensional tissue culture. Two established human colon adenocarcinoma cell lines, HT-29, an undifferentiated, and HT-29KM, a stable, moderately differentiated subline of HT-29, were grown in new tissue culture bioreactors called Rotating-Wall Vessels (RWVs). RWVs are used in conjunction with multicellular cocultivation to develop a unique in vitro tissue modeling system. Cells were cultivated on Cytodex-3 microcarrier beads, with and without mixed normal human colonic fibroblasts, which served as the mesenchymal layer. Culture of the tumor lines in the absence of fibroblasts produced spheroidlike growth and minimal differentiation. In contrast, when tumor lines were co-cultivated with normal colonic fibroblasts, initial growth was confined to the fibroblast population until the microcarriers were covered. The tumor cells then commenced proliferation at an accelerated rate, organizing themselves into three-dimensional tissue masses that achieved 1.0- to 1.5-cm diameters. The masses displayed glandular structures, apical and internal glandular microvilli, tight intercellular junctions, desmosomes, cellular polarity, sinusoid development, internalized mucin, and structural organization akin to normal colon crypt development. Differentiated samples were subjected to transmission and scanning electron microscopy and histologic analysis, revealing embryoniclike mesenchymal cells lining the areas around the growth matrices. Necrosis was minimal throughout the tissue masses. These data suggest that the RWV affords a new model for investigation and isolation of growth, regulatory, and structural processes within neoplastic and normal tissue.

Colonic Neoplasms/pathology↗

Experiments with osteoblasts cultured under hypergravity conditions

To understand further the role of gravity in osteoblast attachment, osteoblasts were subjected to hypergravity conditions in vitro. Scanning electron microscopy of all confluent coverslips from FPA units show that the number of attached osteoblasts was similar among gravitational levels and growth durations (~90 cells/microscopic field). Specifically, confluent 1.0 G control cultures contained an average of 91 +/- 8 cells/field, 3.3 G samples had 88 +/- 8 cells/field, and 4.0 G cultures averaged 90 +/- 7 cells/field. The sparsely plated cultures assessed by immunohistochemistry also had similar numbers of cells at each time point (l.0 G was similar to 3.3 and 4.0 G), but cell number changed from one time point to the next as those cells proliferated. Immunohistochemistry of centrifuged samples showed an increase in number (up to 160% increase) and thickness (up to 49% increase) of actin fibers, a decrease in intensity of fibronectin fluorescence (18-23% decrease) and an increase in number of vinculin bulbs (202-374% increase in number of vinculin bulbs/area). While hypergravity exposure did not alter the number of attached osteoblasts, it did result in altered actin, fibronectin, and vinculin elements, changing some aspects of osteoblast- substrate adhesion.

NASA Program Fundamental Space Biology↗

Response Of Mineralizing And Non-Mineralizing Bone Cells To Fluid Flow: An In Vitro Model For Mechanotransruction

Humans reach peak bone mass at age 30. After this point, we lose 1 to 2 percent of bone mass each decade. In the microgravity environment of space, astronauts lose bone mass at an accelerated rate of 1 to 2 percent each month. When astronauts travel to Mars, they may be in space for as long as 3 years. During this time, they may lose about half of their bone mass from weight-bearing bones. This loss may be irreversible. The drastic loss in bone that astronauts experience in space makes them much more vulnerable to fractures. In addition, the corresponding removal of calcium from bone results in higher levels of calcium in the blood, which increases the risk of developing kidney stones. Currently, studies are being conducted which investigate factors governing bone adaptation and mechanotransduction. Bone is constantly adapting in response to mechanical stimuli. Increased mechanical loading stimulates bone formation and suppresses bone resorption. Reduction in mechanical loading caused by bedrest, disuse, or microgravity results in decreased bone formation and possibly increased bone resorption. Osteoblasts and osteoclasts are the two main cell types that participate in bone remodeling. Osteoblasts are anabolic (bone-forming) cells and osteoclasts are catabolic (bone-resorbing) cells. In microgravity, the activity of osteoblasts slows down and the activity of osteoclasts may speed up, causing a loss of bone density. Mechanotransduction, the molecular mechanism by which mechanical stimuli are converted to biochemical signals, is not yet understood. Exposure of cells to fluid flow imposes a shear stress on the cells. Several studies have shown that the shear stress that results from fluid flow induces a cellular response similar to that induced by mechanical loading. Thus, fluid flow can be used as an in vitro model to simulate the mechanical stress that bone cells experience in vivo. Previous in vitro studies have shown that fluid flow induces several responses in osteoblasts, including increased proliferation, osteoblastic differentiation, alkaline phosphatase activity, and production of nitric oxide, prostaglandins, and osteopontin. Several proteins have been implicated in osteoblastic mechanotransduction including Bone Morphogenetic Protein-2 (BMP-2), parathyroid hormone, 1,25-dihydroxyvitamin D3 receptor, osteopontin (OPN), osteoprotegerin (OPG), and alkaline phosphatase (AP). We will characterize relative levels of each protein in mineralizing or non-mineralizing MC3T3 osteoblastic cells that have been exposed to fluid flow compared to non-fluid flow using immunofluorescent staining and two- photon laser microscopy as well as western blotting. Because calcium-mediated pathways are important in osteoblastic signaling, we will transfect MC3T3 cells with cameleon probes for Ca2+ containing YFP and CFP. Results will be analyzed using FRET/FLIM to study differential release of intracellular Ca(2+) in response to fluid flow and conditions inducing matrix mineralization. In addition, we plan to conduct several microarray experiments to determine differential gene expression in MC3T3 cells in response to fluid flow and conditions inducing mineralization.

Makuch, Lauren A.↗

Environmental impacts on the developing CNS: CD15, NCAM-L1, and GFAP expression in rat neonates exposed to hypergravity

We have previously reported that the developing rat cerebellum is affected by hypergravity exposure. The effect is observed during a period of both granule and glial cell proliferation and neuronal migration in the cerebellum and coincides with changes in thyroid hormone levels. The present study begins to address the molecular mechanisms involved in the cerebellar response to hypergravity. Specifically, the study focuses on the expression of cerebellar proteins that are known to be directly involved in cell-cell interactions [protein expressing 3-fucosyl-N-acetyl-lactosamine antigen (CD15), neuronal cell adhesion molecule (NCAM-L1)] and those that affect cell-cell interactions indirectly [glial fibrillary acidic protein (GFAP)] in rat neonates exposed to centrifuge-produced hypergravity. Cerebellar mass and protein expression in rat neonates exposed to hypergravity (1.5 G) from gestational day (G) 11 to postnatal day (P) 30 were compared at one of six time points between P6 and P30 against rat neonates developing under normal gravity. Proteins were analyzed by quantitative western blots of cerebellar homogenates prepared from male or female neonates. Cerebellar size was most clearly reduced in male neonates on P6 and in female neonates on P9, with a significant gender difference; differences in cerebellar mass remained significant even when change in total body mass was factored in. Densitometric analysis of western blots revealed both quantitative and temporal changes in the expression of selected cerebellar proteins that coincided with changes in cerebellar mass and were gender-specific. In fact, our data indicated certain significant differences even between male and female control animals. A maximal decrease in expression of CD15 was observed in HG females on P9, coinciding with maximal change in their cerebellar mass. A shift in the time-course of NCAM-L1 expression resulted in a significant increase in NCAM-L1 in HG males on P18, an isolated time at which cerebellar mass does not significantly differ between HG and SC neonates. A maximal decrease in expression of GFAP was observed in HG males on P6, coinciding with maximal change in their cerebellar mass. Altered expression of cerebellar proteins is likely to affect a number of developmental processes and contribute to the structural and functional alterations seen in the CNS developing under altered gravity. Our data suggest that both cerebellar development and its response to gravitational manipulations differ in males and females. c2004 COSPAR. Published by Elsevier Ltd. All rights reserved.

NASA Program Fundamental Space Biology↗

Selection of an Alternate Biocide for the ISS Internal Thermal Control System Coolant, Phase 2

The ISS (International Space Station) ITCS (Internal Thermal Control System) includes two internal coolant loops that utilize an aqueous based coolant for heat transfer. A silver salt biocide had previously been utilized as an additive in the coolant formulation to control the growth and proliferation of microorganisms within the coolant loops. Ground-based and in-flight testing demonstrated that the silver salt was rapidly depleted, and did not act as an effective long-term biocide. Efforts to select an optimal alternate biocide for the ITCS coolant application have been underway and are now in the final stages. An extensive evaluation of biocides was conducted to down-select to several candidates for test trials and was reported on previously. Criteria for that down-select included: the need for safe, non-intrusive implementation and operation in a functioning system; the ability to control existing planktonic and biofilm residing microorganisms; a negligible impact on system-wetted materials of construction; and a negligible reactivity with existing coolant additives. Candidate testing to provide data for the selection of an optimal alternate biocide is now in the final stages. That testing has included rapid biocide effectiveness screening using Biolog MT2 plates to determine minimum inhibitory concentration (amount that will inhibit visible growth of microorganisms), time kill studies to determine the exposure time required to completely eliminate organism growth, materials compatibility exposure evaluations, coolant compatibility studies, and bench-top simulated coolant testing. This paper reports the current status of the effort to select an alternate biocide for the ISS ITCS coolant. The results of various test results to select the optimal candidate are presented.

Wilson, Mark E.↗

Protective effect of apigenin on radiation-induced chromosomal damage in human lymphocytes

The potential use of flavonoids as a radioprotector is of increasing interest because of their high antioxidant activity and abundance in the diet. The aim of this study is to examine genotoxic and radioprotective effects of one of the most common flavonoids, apigenin, on radiation-induced chromosome aberrations in human lymphocytes. The cytokinesis-block micronucleus (CBMN) assay was used to evaluate such effects of apigenin. Blood samples were collected from two non-smoking healthy male volunteers who had no history of previous exposure to other clastogenic agents. Isolated lymphocytes were cultured. There were two tubes per concentration for all treatments. To evaluate the genotoxicity of apigenin, cells were first treated with different concentrations of apigenin (0, 2.5, 5, 10 and 25 microg/mL) at 24 h after culture initiation, followed by cytochalasin-B (Cyt-B) treatment (3 microg/mL) and cell harvest at 44 and 72 h, respectively. Secondly, to investigate the radioprotective effect, cell cultures were exposed to different concentrations of apigenin as described above for 30 min before being irradiated to 2 Gy of 137Cs gamma rays (at a dose rate of 0.75 Gy/min). In all instances, the frequency of MN was scored in binucleated (BN) cells. The nuclear proliferation index also was calculated. We did not detect an increase in the frequency of MN in non-irradiated human lymphocyte cultures treated with 2.5, 5.0 or 10 microg/mL apigenin; although, we did observe an increase in cultures treated with 25 microg/mL apigenin (the highest concentration of apigenin used in our study). We also observed a significant increase in the frequency of MN in irradiated cells overall; however, the frequency was decreased as the concentration of apigenin increased, suggesting a radioprotective effect. These findings provide a basis for additional studies to help clarify the potential use and benefit of apigenin as a radioprotector.

NASA Discipline Radiation Health↗

Countermeasure for space flight effects on immune system: nutritional nucleotides

Microgravity and its environment have adverse effects on the immune system. Abnormal immune responses observed in microgravity may pose serious consequences, especially for the recent directions of NASA for long-term space missions to Moon, Mars and deep Space exploration. The study of space flight immunology is limited due to relative inaccessibility, difficulty of performing experiments in space, and inadequate provisions in this area in the United States and Russian space programs (Taylor 1993). Microgravity and stress experienced during space flights results in immune system aberration (Taylor 1993). In ground-based mouse models for some of the microgravity effects on the human body, hindlimb unloading (HU) has been reported to cause abnormal cell proliferation and cytokine production (Armstrong et al., 1993, Chapes et al. 1993). In this report, we document that a nutritional nucleotide supplementation as studied in ground-based microgravity analogs, has potential to serve as a countermeasure for the immune dysfunction observed in space travel.

NASA Center JSC↗

Multiple melanocortin receptors are expressed in bone cells

Melanocortin receptors belong to the seven transmembrane domain, G-protein coupled family of receptors. There are five members of this receptor family labeled MC1R-MC5R. These receptors are activated by fragments derived from a larger molecule, proopiomelanocortin (POMC) and include ACTH, alpha beta and gamma-MSH and beta-endorphin. Because of in vitro and in vivo data suggesting direct effects of these POMC molecules on bone and bone turnover, we examined bone and bone derived cells for the presence of the various members of the melanocortin receptor family. We report that the five known melanocortin receptors are expressed to varying degrees in osteoblast-like and osteoclastic cells. POMC fragments increased proliferation and expression of a variety of genes in osteoblastic cells. Furthermore, POMC mRNA was detected in osteoclastic cells. These data demonstrate that POMC-derived peptide hormones acting through high affinity melanocortin receptors have specific effects on bone cells. Thus, in addition to the indirect effects of POMC-derived hormones on bone turnover through their modulation of steroid hormone secretion, POMC fragments may have direct and specific effects on bone cell subpopulations.

NASA Discipline Musculoskeletal↗

Modeling Leadership Styles in Human-Robot Team Dynamics

The recent proliferation of robotic systems in our society has placed questions regarding interaction between humans and intelligent machines at the forefront of robotics research. In response, our research attempts to understand the context in which particular types of interaction optimize efficiency in tasks undertaken by human-robot teams. It is our conjecture that applying previous research results regarding leadership paradigms in human organizations will lead us to a greater understanding of the human-robot interaction space. In doing so, we adapt four leadership styles prevalent in human organizations to human-robot teams. By noting which leadership style is more appropriately suited to what situation, as given by previous research, a mapping is created between the adapted leadership styles and human-robot interaction scenarios-a mapping which will presumably maximize efficiency in task completion for a human-robot team. In this research we test this mapping with two adapted leadership styles: directive and transactional. For testing, we have taken a virtual 3D interface and integrated it with a genetic algorithm for use in &le-operation of a physical robot. By developing team efficiency metrics, we can determine whether this mapping indeed prescribes interaction styles that will maximize efficiency in the teleoperation of a robot.

Cruz, Gerardo E.↗

Layered virus protection for the operations and administrative messaging system

NASA's Deep Space Network (DSN) is critical in supporting the wide variety of operating and plannedunmanned flight projects. For day-to-day operations it relies on email communication between the three Deep Space Communication Complexes (Canberra, Goldstone, Madrid) and NASA's Jet Propulsion Laboratory. The Operations & Administrative Messaging system, based on the Microsoft Windows NTand Exchange platform, provides the infrastructure that is required for reliable, mission-critical messaging. The reliability of this system, however, is threatened by the proliferation of email viruses that continue to spread at alarming rates. A layered approach to email security has been implemented across the DSN to protect against this threat.

IT security antivirus↗

Packaging and Qualification of Mems-Based Space Systems

The number of spacecraft designed and built over the next century will grow exponentially as communication satellite networks proliferate and NASA continues to push towards the development of many microspacecraft to replace its traditional.

communication satellite networks satellite network↗

Diversification and Challenges of Software Engineering Standards

The author poses certain questions in this paper: 'In the future, should there be just one software engineering standards set? If so, how can we work towards that goal? What are the challenges of internationalizing standards?' Based on the author's personal view, the statement of his position is as follows: 'There should NOT be just one set of software engineering standards in the future. At the same time, there should NOT be the proliferation of standards, and the number of sets of standards should be kept to a minimum.It is important to understand the diversification of the areas which are spanned by the software engineering standards.' The author goes on to describe the diversification of processes, the diversification in the national and international character of standards organizations, the diversification of the professional organizations producing standards, the diversification of the types of businesses and industries, and the challenges of internationalizing standards.

software standards international standards softwar↗