Characterization of Physiological Changes in Roots Grown in Spaceflight Conditions: A Comparison of Nutrient Delivery Technologies.

The best strategy for providing food and maintaining the environment on long-duration space missions is a bioregenerative life support system based on the growth of higher plants. Before such a system can be implemented, a better understanding of plant growth in space will have to be achieved. Little is known about the role of gravity-dependent physical processes in normal physiological function. A series of ground-based and spaceflight experiments was conducted to examine root oxygen availability in microgravity nutrient delivery systems. In spaceflight experiments Arabidopsis thaliana (L.) Heynh. plants were analyzed for changes in root medium redox potential and root alcohol dehydrogenase (ADH) activity, localization, and expression. These experiments showed ADH activity and expression increased by 89% and 136% respectively, without any change in localization. Ground experiments demonstrated the increase in ADH activity in spaceflight roots was achieved by a 28% decrease in oxygen availability. Metabolic and growth pattern changes in spaceflight roots suggested that root orientation could be directed toward oxygen (oxytropism). Both gravity sensing and agravitropic Pisum sativum L. roots reoriented toward oxygen when grown in a microrhizotron that maintains an oxygen gradient over the whole range of subambient oxygenation, although the rate of reorientation declined as oxygen concentrations decreased. Ground-based experiments to evaluate technologies for growing plants in space were conducted by comparing root system morphology, ADH activity, nutrient content, and general plant growth parameters of Triticum aestivum L. cv Yecora Rojo and Brassica rapa L. A system that maintains a nutrient solution inside of a porous hydrophilic ceramic tube was compared with two passive substrate systems now used for spaceflight research (solidified agar-gel and phenolic foam media). Plants performed best on the porous tube because the roots grow on the tube surface in contact with air and nutrient solution. Oxygenation is important for normal root function in space, as on earth. Two obstacles restricting development of plant nutrient delivery technologies are a limited knowledge of the requirements of plants in space, and an incomplete understanding of plants in general. Information concerning the changes in oxygenation during spaceflight should allow scientists and engineers to address these problems

A non-invasive method for measuring preimplantation embryo physiology

Author Posting. © Cambridge University Press, 2000. This article is posted here by permission of Cambridge University Press for personal use, not for redistribution. The definitive version was published in Zygote 8 (2000): 15-24, doi:10.1017/S0967199400000782.The physiology of the early embryo may be indicative of embryo vitality and therefore methods for non-invasively monitoring physiological parameters from embryos could improve preimplantation diagnoses. The self-referencing electrophysiological technique is capable of non-invasive measurement of the physiology of individual cells by monitoring the movement of ions and molecules between the cell and the surrounding media. Here we use this technique to monitor gradients of calcium, potassium, oxygen and hydrogen peroxide around individual mouse preimplantation embryos. The calcium-sensitive electrode in self-referencing mode identified a region of elevated calcium concentration ([similar]0.25 pmol) surrounding each embryo. The calcium gradient surrounding embryos was relatively steep, such that the region of elevated calcium extended into the medium only 4 [mu]m from the embryo. By contrast, using an oxygen-sensitive electrode an extensive gradient of reduced dissolved oxygen concentration was measured surrounding the embryo and extended tens of micrometres into the medium. A gradient of neither potassium nor hydrogen peroxide was observed around unperturbed embryos. We also demonstrate that monitoring the physiology of embryos using the self-referencing technique does not compromise their subsequent development. Blastocysts studied with the self-referencing technique implanted and developed to term at the same frequency as did unexamined, control embryos. Therefore, the self-referencing electrode provides a valuable non-invasive technique for studying the physiology and pathophysiology of individual embryos without hindering their subsequent development.A portion of this work was funded by an NIH R21 #RR 12718–02 to D.L.K. and P.J.S.S., KO81099 to D.L.K. and NIH P41 RR01395 to P.J.S.S

Mouse and human islets survive and function after coating by biosilicification

Inorganic materials have properties that can be advantageous in bioencapsulation for cell transplantation. Our aim was to engineer a hybrid inorganic/soft tissue construct by inducing pancreatic islets to grow an inorganic shell. We created pancreatic islets surrounded by porous silica, which has potential application in the immunoprotection of islets in transplantation therapies for type 1 diabetes. The new method takes advantage of the islet capsule surface as a template for silica formation. Mouse and human islets were exposed to medium containing saturating silicic acid levels for 9-15 min. The resulting tissue constructs were then cultured for up to 4 wk under normal conditions. Scanning electron microscopy and energy dispersive X-ray spectroscopy was used to monitor the morphology and elemental composition of the material at the islet surface. A cytokine assay was used to assess biocompatibility with macrophages. Islet survival and function were assessed by confocal microscopy, glucose-stimulated insulin release assays, oxygen flux at the islet surface, expression of key genes by RT-PCR, and syngeneic transplant into diabetic mice

The Science Performance of JWST as Characterized in Commissioning

This paper characterizes the actual science performance of the James Webb Space Telescope (JWST), as determined from the six month commissioning period. We summarize the performance of the spacecraft, telescope, science instruments, and ground system, with an emphasis on differences from pre-launch expectations. Commissioning has made clear that JWST is fully capable of achieving the discoveries for which it was built. Moreover, almost across the board, the science performance of JWST is better than expected; in most cases, JWST will go deeper faster than expected. The telescope and instrument suite have demonstrated the sensitivity, stability, image quality, and spectral range that are necessary to transform our understanding of the cosmos through observations spanning from near-earth asteroids to the most distant galaxies.Comment: 5th version as accepted to PASP; 31 pages, 18 figures; https://iopscience.iop.org/article/10.1088/1538-3873/acb29

Effects of Carbon Nanotube-Tethered Nanosphere Density on Amperometric Biosensing: Simulation and Experiment

Nascent nanofabrication approaches are being applied to reduce electrode feature dimensions from the microscale to the nanoscale, creating biosensors that are capable of working more efficiently at the biomolecular level. The development of nanoscale biosensors has been driven largely by experimental empiricism to date. Consequently, the precise positioning of nanoscale electrode elements is typically neglected, and its impact on biosensor performance is subsequently overlooked. Herein, we present a bottom-up nanoelectrode array fabrication approach that utilizes low-density and horizontally oriented single-walled carbon nanotubes (SWCNTs) as a template for the growth and precise positioning of Pt nanospheres. We further develop a computational model to optimize the nanosphere spatial arrangement and elucidate the trade-offs among kinetics, mass transport, and charge transport in an enzymatic biosensing scenario. Optimized model variables and experimental results confirm that tightly packed Pt nanosphere/SWCNT nanobands outperform low-density Pt nanosphere/SWCNT arrays in enzymatic glucose sensing. These computational and experimental results demonstrate the profound impact of nanoparticle placement on biosensor performance. This integration of bottom-up nanoelectrode array templating with analysis-informed design produces a foundation for controlling and optimizing nanotechnology-based electrochemical biosensor performance

Oxidative phosphorylation-dependent and -independent oxygen consumption by individual preimplantation mouse embryos

The self-referencing electrode technique was employed to noninvasively measure gradients of dissolved oxygen in the medium immediately surrounding developing mouse embryos and, thereby, characterized changes in oxygen consumption and utilization during development. A gradient of depleted oxygen surrounded each embryo and could be detected >50 microm from the embryo. Blastocysts depleted the surrounding medium of 0.6+/-0.1 microM of oxygen, whereas early cleavage stage embryos depleted the medium of only 0.3+/-0.1 microM of oxygen, suggesting a twofold increase in oxygen consumption at the blastocyst stage. Mitochondrial oxidative phosphorylation (OXPHOS) accounted for 60-70% of the oxygen consumed by blastocysts, while it accounted for only 30% of the total oxygen consumed by cleavage-stage embryos. The amount of oxygen consumed by non-OXPHOS mechanisms remained relatively constant throughout preimplantation development. By contrast, the amount of oxygen consumed by OXPHOS in blastocysts is greater than that consumed by OXPHOS in cleavage-stage embryos. The amount of oxygen consumed by one-cell embryos was modulated by the absence of pyruvate from the culture medium. Treatment of one-cell embryos and blastocysts with diamide, an agent known to induce cell death in embryos, resulted in a decline in oxygen consumption, such that the medium surrounding dying embryos was not as depleted of oxygen as that surrounding untreated control embryos. Together these results validate the self-referencing electrode technique for analyzing oxygen consumption and utilization by preimplantation embryos and demonstrate that changes in oxygen consumption accompany important physiological events, such as development, response to medium metabolites, or cell deat

Lab-on-a-chip approaches for space-biology research

Lab-on-a-chip (LOC) systems with electrochemical sensing capability can provide real-time physiological measurements in spaceflight environments. They are easily miniaturized and integrated with existing space hardware systems. To reduce crew time during spaceflight research, the systems can be made autonomous and simple to use. Research and development of electrochemical-sensing LOC systems are still in progress for fundamental space-biology research in microgravity. Ion-selective electrodes as electrochemical sensors are miniaturized in an all-solid-state format for easier packaging and handling. The design, fabrication, and application of these sensors are discussed, with examples from those developed at the Physiological Sensing Facility (PSF) at Purdue University. The objective of this paper is not to provide an exhaustive review of current LOC systems, but to describe research developments made for the purpose of conducting physiological measurements in microgravity with examples of patents that support space missions

A comparative study of enzyme immobilization strategies for multi-walled carbon nanotube glucose biosensors

This work addresses the comparison of different strategies for improving biosensor performance using nanomaterials. Glucose biosensors based on commonly applied enzyme immobilization approaches, including sol-gel encapsulation approaches and glutaraldehyde cross-linking strategies, were studied in the presence and absence of multi-walled carbon nanotubes (MWNTs). Although direct comparison of design parameters such as linear range and sensitivity is intuitive, this comparison alone is not an accurate indicator of biosensor efficacy, due to the wide range of electrodes and nanomaterials available for use in current biosensor designs. We proposed a comparative protocol which considers both the active area available for transduction following nanomaterial deposition and the sensitivity. Based on the protocol, when no nanomaterials were involved, TEOS/GOx biosensors exhibited the highest efficacy, followed by BSA/GA/GOx and TMOS/GOx biosensors. A novel biosensor containing carboxylated MWNTs modified with glucose oxidase and an overlying TMOS layer demonstrated optimum efficacy in terms of enhanced current density (18.3 +/- 0.5 mu A mM(-1) cm(-2)), linear range (0.0037-12 mM), detection limit (3.7 mu M), coefficient of variation (2%), response time (less than 8 s), and stability/selectivity/reproducibility. H(2)O(2) response tests demonstrated that the most possible reason for the performance enhancement was an increased enzyme loading. This design is an excellent platform for versatile biosensing applications

Control of ascorbic acid efflux in rat luteal cells: role of intracellular calcium and oxygen radicals

In luteal cells, prostaglandin (PG)F2a mobilizes intracellular calcium concentration ([Ca]i), generates reactive oxygen species (ROS), depletes ascorbic acid (AA) levels, inhibits steroidogenesis, and ultimately induces cell death. We investigated the hypothesis that [Ca]i mobilization stimulates ROS, which results in depletion of cellular AA in rat luteal cells. We used a self-referencing AA-selective electrode that noninvasively measures AA flux at the extended boundary layer of single cells and fluorescence microscopy with fura 2 and dichlorofluorescein diacetate (DCF-DA) to measure [Ca]i and ROS, respectively. Menadione, a generator of intracellular superoxide radical (O2-), PGF2a, and calcium ionophore were shown to increase [Ca]i and stimulate intracellular ROS. With calcium ionophore and PGF2a, but not menadione, the generation of ROS was dependent on extracellular calcium influx. In unstimulated cells there was a net efflux of AA of 121.5 +/- 20.3 fmol x cm-1 x s-1 (mean +/- SE, n = 8), but in the absence of extracellular calcium the efflux was significantly reduced (10.3 +/- 4.9 fmol x cm-1 x s-1; n = 5, P < 0.05). PGF2a and menadione stimulated AA efflux, but calcium ionophore had no significant effect. These data suggest two AA regulatory mechanisms: Under basal conditions, AA efflux is calcium dependent and may represent recycling and maintenance of an antioxidant AA gradient at the plasma membrane. Under luteolytic hormone and/or oxidative stress, AA efflux is stimulated that is independent of extracellular calcium influx or generation of ROS. Although site-specific mobilization of calcium pools and ROS cannot be ruled out, the release of AA by PGF2a-stimulated luteal cells may occur through other signaling pathways

Nanostructuring Platinum Nanoparticles on Multilayered Graphene Petal Nanosheets for Electrochemical Biosensing

Hybridization of nanoscale metals and carbon nanotubes into composite nanomaterials has produced some of the best-performing sensors to date. The challenge remains to develop scalable nanofabrication methods that are amenable to the development of sensors with broad sensing ranges. A scalable nanostructured biosensor based on multilayered graphene petal nanosheets (MGPNs), Pt nanoparticles, and a biorecognition element (glucose oxidase) is presented. The combination of zero-dimensional nanoparticles on a two-dimensional support that is arrayed in the third dimension creates a sensor platform with exceptional characteristics. The versatility of the biosensor platform is demonstrated by altering biosensor performance (i.e., sensitivity, detection limit, and linear sensing range) through changing the size, density, and morphology of electrodeposited Pt nanoparticles on the MGPNs. This work enables a robust sensor design that demonstrates exceptional performance with enhanced glucose sensitivity (0.3 mu M detection limit, 0.0150 mM linear sensing range), a long stable shelf-life (\u3e1 month), and a high selectivity over electroactive, interfering species commonly found in human serum samples