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Effect of 14 days of bed rest on urine metabolite excretion and plasma enzyme levels

After 1 week of ambulatory base-line measurement, a group of 8 men 19-26 years of age remained continuously recumbent for 14 days. Studies were continued for 1 week following the prolonged recumbency. Urine excretion rates for a number of constituents were determined 2 days before bed rest, on day 14 of bed rest, and day 6 after bed rest. Blood plasma samples were also obtained at these times, and analyzed for several enzymes. On day 14 of bed rest significant increases were observed in urine excretion of total osmotically-active substances, magnesium, calcium, phosphate, creatinine, hydroxyproline, and 17-OH corticosteroids. A decrease occurred in urinary glucose excretion. Plasma levels of alkaline phosphatase and LDH-3 were depressed, while plasma GPT was elevated. Many of these changes persisted on day 6 after bed rest, and are interpreted as concomitants of the disuse atrophy of the musculoskeletal system that characterizes prolonged bed rest and weightlessness.

Pace, N.↗

Results of the TTF-TCNQ and the calcium carbonate crystallization on the Long Duration Exposure Facility

Experiment A0139A on the Long Duration Exposure Facility (LDEF) carried four large containers into orbit five years with crystal growth solutions for lead sulfide, calcium carbonate, and TTF-TCNQ. Although temperature data was lost, the experimental program had been working since the valves in all containers had been opened. All four experiments produced crystals of varying quality. The calcium carbonate crystals had the best appearance. The TTF-TCNQ crystals were packed together near the valve openings of the container. When taken apart, the single crystals showed some unusual morphological properties. X ray investigations as well as conductivity measurements on long duration space grown TTF-TCNQ crystals will be presented. Comparisons will be made with our previous space solution growth experiments. The TTF-TCNQ crystals are no longer of the highest interest, so this activity has been terminated in favor of calcium carbonate and calcium phosphate crystallizations.

Nielsen, Kjeld Flemming↗

Follow up on the crystal growth experiments of the LDEF

The 4 solution growth experiments on the LDEF were presented thoroughly elsewhere. The CaCO3-experiment, and to a certain extent also the TTF-TCNQ-experiments yielded useful results. In Jan. 1992, the next series of solution growth experiments were sent to ESA for shipment to KSC. As on the LDEF, the SGF (solution growth facility) of the EURECA-1 contains 4 large experiments. From the beginning, the planning and developments were introduced. Still, the basic concept was maintained, and the CaCO3-experiment, that showed the best results on the LDEF, will now be repeated with improved technology and in larger scale on the EURECA-1. The contents of the 4 SGF experiments are as follows: (1) growth of calcium-carbonate crystals; (2) formation and transformation of tri-calcium-phosphate; (3) growth of zeolite crystals; and (4) soret coefficient measurements (diffusion). The scientific background for the choice of experiments and the major improvements of the SGF are reviewed. Furthermore, some ideas on basic microgravity solution growth experimentation from ESA's newly established EWG (Expert Working Group) on solution growth are reported.

Nielsen, K. F.↗

Slow-release fertilizer

A synthetic apatite containing agronutrients and a method for making the apatite are disclosed. The apatite comprises crystalline calcium phosphate having agronutrients dispersed in the crystalline structure. The agronutrients can comprise potassium, magnesium, sulfur, iron, manganese, molybdenum, chlorine, boron, copper and zinc in amounts suited for plant growth. The apatite can optionally comprise a carbonate and/or silicon solubility control agent. The agronutrients are released slowly as the apatite dissolves.

Ming, Douglas W.↗

Active synthetic soil

A synthetic soil/fertilizer for horticultural application having all the agronutrients essential for plant growth is disclosed. The soil comprises a synthetic apatite fertilizer having sulfur, magnesium, and micronutrients dispersed in a calcium phosphate matrix, a zeolite cation exchange medium saturated with a charge of potassium and nitrogen cations, and an optional pH buffer. Moisture dissolves the apatite and mobilizes the nutrient elements from the apatite matrix and the zeolite charge sites.

Ming, Douglas W.↗

Evidence for Life on Earth before 3,800 Million Years Ago

It is unknown when life first appeared on Earth. The earliest known microfossils (approx. 3,500 Myr before present) are structurally complex, and if it is assumed that the associated organisms required a long time to develop this degree of complexity, then the existence of life much earlier than this can be argued. But the known examples of crustal rocks older than approx. 3,500 Myr have experienced intense metamorphism, which would have obliterated any fragile microfossils contained therein. It is therefore necessary to search for geochemical evidence of past biotic activity that has been preserved within minerals that are resistant to metamorphism. Here we report ion-microprobe measurements of the carbon-isotope composition of carbonaceous inclusions within grains of apatite (basic calcium phosphate) from the oldest known sediment sequences a approx. 3,800 Myr-old banded iron formation from the Isua supracrustal belt, West Greenland, and a similar formation from the nearby Akilia island that is possibly older than 3,850 Myr. The carbon in the carbonaceous inclusions is isotopically light, indicative of biological activity; no known abiotic process can explain the data. Unless some unknown abiotic process exists which is able both to create such isotopically light carbon and then selectively incorporate it into apatite grains, our results provide evidence for the emergence of life on Earth by at least 3,800 Myr before present.

Mojzsis, S. J.↗

A Structural and Molecular Approach for the Study Biomarkers

Investigation of the nucleation and growth of crystals in both abiotic and biotic systems is critical to seemingly diverse disciplines of geology, biology, environmental science, and astrobiology. While there are abundant studies devoted to the determination of the structure and composition of inorganic crystals, as well as to the development of thermodynamic and kinetic models, it is only recently that research efforts have been directed towards understanding mineralization in biological systems (i.e., biomineralization). Biomineralization refers to the processes by which living organisms form inorganic solids. Studies of the processes of biomineralization under low temperature aqueous conditions have focused primarily on magnetite forming bacteria and shell forming marine organisms. Many of the biological building materials consist of inorganic minerals (calcium carbonate, calcium phosphate, silica or iron oxide) intricately combined with organic polymers (like proteins). More recently, efforts have been undertaken to explore the nature of biological activities in ancient rocks. In the absence of well-preserved microorganisms or genetic material required for the polmerase chain reaction (PCR) method in molecular phylogenetic studies, using biominerals as biomarkers offers an alternative approach for the recognition of biogenic activity in both terrestrial and extraterrestrial environments. The primary driving force in biomineralization is the interaction between organic and inorganic phases. Thus, the investigation of the ultrastructure and the nature of reactions at the molecular level occurring at the interface between inorganic and organic phases is essential to understanding the processes leading to the nucleation and growth of crystals. It is recognized that crystal surfaces can serve as the substrate for the organization of organic molecules that lead to the formation of polymers and other complex organic molecules, and in discussions of the origins of life, is referred to as organic synthesis on mineral surfaces. Furthermore, it is suggested that the interaction between mineral surfaces and simple organic molecules resulted in the formation of amino acids, RNA, and perhaps other more complex molecules such as proteins. On the other hand, in natural systems, it is recognized that functional groups on cell walls or membranes of microorganisms serve as sites of nucleation and crystallization. The precise replication of biominerals with controlled structure, morphology, size and texture is not confined to higher organisms as it also occurs in primitive prokaryotic cells such as magnetotactic bacteria and cyanobacteria. This suggests that the principal strategies of biomineralization were established early on in the evolutionary history of organisms. It is critical, therefore, to search for common mechanisms within diverse biological systems. One such common factor is the capability for organization and self-assembly. Organic macromolecules such as proteins and lipids can aggregate and polymerize forming membranes or extracellular matrix. At the organic-inorganic interface, several factors such as lattice geometry, polarity, stereochemistry and topography may act in concert to control nucleation and growth of crystals. Although several models have been proposed that discuss the significance of these factors for biomineralization, no comprehensive experimental data are available. In contrast to crystallization in exclusively inorganic systems, the kinetics of reaction and structural relationships between organic and inorganic phases in biominerals or biomimetic material is poorly understood. For example, it is not clear if the concept of epitactic growth (geometrical matching of unit cells at the interface of a secondary crystal growing on a primary crystal) applies to organic-inorganic systems. In contrast to inorganic templates that often have a smooth and rigid surface that promotes epitactic growth, biological substrates are usually rough and result in a large degree of mismatch. It is apparent that factors controlling the reaction at the crystal-matrix interface are strongly dependent upon the nature of the substrate. Therefore, characterization of the assembled organic surface and surface structure of the inorganic phase is crucial to understanding the processes of biomineralization. The focus of our research is the investigation of the processes leading to the nucleation and growth of crystals on both natural and synthetic systems through an interdisciplinary approach that integrates molecular biology, morphology and mineralogy using advanced preparation and analytical techniques. We have studied run-products, particularly magnetite, siderite and other carbonates, that resulted from extracellular biomineralization by extremophiles isolated from a variety of extreme environments ranging from permafrost to hydrothermal vent systems. The results of this study are critical to recognizing biomarkers in terrestrial and extraterrestrial environments.

Thomas-Keprta, Kathie↗

Dissolution kinetics of iron-, manganese-, and copper-containing synthetic hydroxyapatites

Micronutrient-substituted synthetic hydroxyapatite (SHA) is being evaluated by the National Aeronautics and Space Administration's (NASA) Advanced Life Support (ALS) Program for crop production on long-duration human missions to the International Space Station or for future Lunar or Martian outposts. The stirred-flow technique was utilized to characterize Ca, P, Fe, Mn, and Cu release characteristics from Fe-, Mn-, and Cu-containing SHA in deionized (DI) water, citric acid, and diethylene-triamine-pentaacetic acid (DTPA). Initially, Ca and P release rates decreased rapidly with time and were controlled by a non-SHA calcium phosphate phase(s) with low Ca/P solution molar ratios (0.91-1.51) relative to solid SHA ratios (1.56-1.64). At later times, Ca/P solution molar ratios (1.47-1.79) were near solid SHA ratios and release rates decreased slowly indicating that SHA controlled Ca and P release. Substituted SHA materials had faster dissolution rates relative to unsubstituted SHA. The initial metal release rate order was Mn >> Cu > Fe which followed metal-oxide/phosphate solubility suggesting that poorly crystalline metal-oxides/phosphates were dominating metal release. Similar metal release rates for all substituted SHA (approximately 0.01 cmol kg-1 min-1) at the end of the DTPA experiment indicated that SHA dissolution was supplying the metals into solution and that poorly crystalline metal-oxide/phosphates were not controlling metal release. Results indicate that non-SHA Ca-phosphate phases and poorly crystalline metal-oxide/phosphates will contribute Ca, P, and metals. After these phases have dissolved, substituted SHA will be the source of Ca, P, and metals for plants.

NASA Discipline Life Support Systems↗

Persistent Seroconversion after Accidental Eye Exposure to Calcifying Nanoparticles

Biosafety of nanomaterials has attracted much attention recently. We report here a case where accidental human eye exposure to biogenic nanosized calcium phosphate in the form of calcifying nanoparticles (CNP) raised a strong IgG immune response against proteins carried by CNP. The antibody titer has persisted over ten years at the high level. The IgG was detected by ELISA using CNPs propagated in media containing bovine and human serum as antigen. The exposure incident occurred to a woman scientist (WS) at a research laboratory in Finland at 1993. CNP, also termed "nanobacteria", is a unique self-replicating agent that has not been fully characterized and no data on biohazards were available at that time. Before the accident, her serum samples were negative for both CNP antigen and anti-CNP antibody using specific ELISA tests (Nanobac Oy, Kuopio, Finland). The accident occurred while WS was harvesting CNP cultures. Due to a high pressure in pipetting, CNP pellet splashed into her right eye. Both eyes were immediately washed with water and saline. The following days there was irritation and redness in the right eye. These symptoms disappeared within two weeks without any treatment. Three months after the accident, blood and urine samples of WS were tested for CNP cultures (2), CNP-specific ELISA tests, and blood cell counts. Blood cell counts were normal, CNP antigen and culture tests were negative. A high IgG anti-CNP antibody titer was detected (see Figure). The antibodies of this person have been used thereafter as positive control and standard in ELISA manufacturing (Nano-Sero IgG ELISA, Nanobac Oy, Kuopio, Finland).

Ciftcioglu, Neva↗

An Additional Potential Factor for Kidney Stone Formation during Space Flights: Calcifying Nanoparticles (Nanobacteria): A Case Report

Spaceflight-induced microgravity appears to be a risk factor for the development of urinary calculi due to skeletal calcium liberation and other undefined factors, resulting in stone disease in crewmembers during and after spaceflight. Calcifying nanoparticles, or nanobacteria, reproduce at a more rapid rate in simulated microgravity conditions and create external shells of calcium phosphate in the form of apatite. The questions arises whether calcifying nanoparticles are niduses for calculi and contribute to the development of clinical stone disease in humans, who possess environmental factors predisposing to the development of urinary calculi and potentially impaired immunological defenses during spaceflight. A case of a urinary calculus passed from an astronaut post-flight with morphological characteristics of calcifying nanoparticles and staining positive for a calcifying nanoparticle unique antigen, is presented.

Jones, Jeffrey A.↗

Active synthetic soil

A synthetic soil/fertilizer for horticultural application having all the agronutrients essential for plant growth is disclosed. The soil comprises a synthetic apatite fertilizer having sulfur, magnesium and micronutrients dispersed in a calcium phosphate matrix, a zeolite cation exchange medium saturated with a charge of potassium and nitrogen cations, and an optional pH buffer. Moisture dissolves the apatite and mobilizes the nutrient elements from the apatite matrix and the zeolite charge sites.

Ming, Douglas W.↗

Slow-release fertilizer

A synthetic apatite containing agronutrients and a method for making the apatite are disclosed. The apatite comprises crystalline calcium phosphate having agronutrients dispersed in the crystalline structure. The agronutrients can comprise potassium, magnesium, sulfur, iron, manganese, molybdenum, chlorine, boron, copper and zinc in amounts suited for plant growth. The apatite can optionally comprise a carbonate and/or silicon solubility control agent. The agronutrients are released slowly as the apatite dissolves.

Ming, Douglas W.↗

Plant Growth Experiments in Zeoponic Substrates: Applications for Advanced Life Support Systems

A zeoponic plant-growth system is defined as the cultivation of plants in artificial soils, which have zeolites as a major component (Allen and Ming, 1995). Zeolites are crystalline, hydrated aluminosilicate minerals that have the ability to exchange constituent cations without major change of the mineral structure. Recently, zeoponic systems developed at the National Aeronautics and Space Administration (NASA) slowly release some (Allen et at., 1995) or all of the essential plant-growth nutrients (Ming et at., 1995). These systems have NH4- and K-exchanged clinoptilolite (a natural zeolite) and either natural or synthetic apatite (a calcium phosphate mineral). For the natural apatite system, Ca and P were made available to the plant by the dissolution of apatite. Potassium and NH4-N were made available by ion-exchange reactions involving Ca(2+) from apatite dissolution and K(+) and NH4(+) on zeolitic exchange sites. In addition to NH4-N, K, Ca, and P, the synthetic apatite system also supplied Mg, S, and other micronutrients during dissolution (Figure 1). The overall objective of this research task is to develop zeoponic substrates wherein all plant growth nutrients are supplied by the plant growth medium for several growth seasons with only the addition of water. The substrate is being developed for plant growth in Advanced Life Support (ALS) testbeds (i.e., BioPLEX) and microgravity plant growth experiments. Zeoponic substrates have been used for plant growth experiments on two Space Shuttle flight experiments (STS-60; STS-63; Morrow et aI., 1995). These substrates may be ideally suited for plant growth experiments on the International Space Station and applications in ALS testbeds. However, there are several issues that need to be resolved before zeoponics will be the choice substrate for plant growth experiments in space. The objective of this paper is to provide an overview on recent research directed toward the refinement of zeoponic plant growth substrates.

Ming, Douglas W.↗

Ex Vivo Growth of Bioengineered Ligaments and Other Tissues

A method of growing bioengineered tissues for use in surgical replacement of damaged anterior cruciate ligaments has been invented. An anterior cruciate ligament is one of two ligaments (the other being the posterior cruciate ligament) that cross in the middle of a knee joint and act to prevent the bones in the knee from sliding forward and backward relative to each other. Anterior cruciate ligaments are frequently torn in sports injuries and traffic accidents, resulting in pain and severe limitations on mobility. By making it possible to grow replacement anterior cruciate ligaments that structurally and functionally resemble natural ones more closely than do totally synthetic replacements, the method could create new opportunities for full or nearly full restoration of functionality in injured knees. The method is also adaptable to the growth of bioengineered replacements for other ligaments (e.g., other knee ligaments as well as those in the hands, wrists, and elbows) and to the production of tissues other than ligaments, including cartilage, bones, muscles, and blood vessels. The method is based on the finding that the histomorphological properties of a bioengineered tissue grown in vitro from pluripotent cells within a matrix are affected by the direct application of mechanical force to the matrix during growth generation. This finding provides important new insights into the relationships among mechanical stress, biochemical and cell-immobilization methods, and cell differentiation, and is applicable to the production of the variety of tissues mentioned above. Moreover, this finding can be generalized to nonmechanical (e.g., chemical and electromagnetic) stimuli that are experienced in vivo by tissues of interest and, hence, the method can be modified to incorporate such stimuli in the ex vivo growth of replacements for the various tissues mentioned above. In this method, a three-dimensional matrix made of a suitable material is seeded with pluripotent stem cells. The patient s bone-marrow stromal cells are preferably used as the pluripotent cells in this method. Suitable matrix materials are materials to which cells can adhere for example, collagen type I. The seeded matrix is attached to anchors at opposite ends and then the cells in the matrix are cultured under conditions appropriate for the growth and regeneration of cells. Suitable anchor materials are materials to which the matrix can attach; examples include demineralized bone and Goinopra coral that has been treated to convert its calcium carbonate to calcium phosphate.

Altman, Gregory↗

Experimentally Shocked and Altered Basalt: Laboratory Analogs for Calibration of Mars Remote Sensing and In Situ Data

Calcium phosphate (likely chloroapatite) is formed in the alteration experiments and is more abundant in the altered and shocked sample probably due to increased surface area exposed to alteration fluids resulting from shock damage in the form of both brittle and structural deformation to the starting material (Figs 1 & 3). Apatite forms in basic conditions so the closed system alteration experiment must be buffered by the basalt starting material to create a fluid chemistry environment evolving from neutral at the start to alkaline after 21 days at 160 degrees Centigrade. Plagioclase feldspar in the unshocked sample (Fig. 2) has undergone a solid-state transformation to maskelynite, a disordered phase that is not manifest in the X-ray diffraction pattern of the shocked sample (Fig.4). Olivine and ulvospinel that are present in the starting material can be detected by X-ray diffraction in the shocked and altered sample (Fig. 4). Tungsten from the sample holder used in the shock experiments dominates the X-ray diffraction pattern of the shocked and altered sample (Fig. 4). Samples were weighed after the alteration experiments to determine mass loss and predict the amount of material available for the planned analyses from the shock experiments. Within the constraints of these experiments, mass loss is negligible. The samples will next be characterized by Moessbauer and Vis-Near Infrared spectroscopy, the results of which will be compared to the Mars Exploration Rovers and Mars Reconnaissance Orbiter data sets respectively.

Bell, M. S.↗

Nanobacteria: Fact or Fiction? Characteristics, Detection and Medical Importance of Novel Self-Replicating, Calcifying Nanoparticles

There is some debate in microbiology as to whether Nanobacteria (NB) are alive. This paper reviews some aspects of NB. In summary, Nanobacteria is a perfect model for studying biogenic mineralization/calcification because NB a) are self-replicating particles and have less complicated metabolic pathways b) accumulate calcium and phosphate under physiological conditions, c)produce a calcium phosphate mineral similar to bone, d) exist in physical conditions (pH, gravity, temperature, etc) that are easy to manipulate, and which can be replicated for the physiological model.

Ciftcioglu, Neva↗

A potential cause for kidney stone formation during space flights: enhanced growth of nanobacteria in microgravity

BACKGROUND: Although some information is available regarding the cellular/molecular changes in immune system exposed to microgravity, little is known about the reasons of the increase in the kidney stone formation in astronauts during and/or after long duration missions at zero gravity (0 g). In our earlier studies, we have assessed a unique agent, nanobacteria (NB), in kidney stones and hypothesized that NB have an active role in calcium phosphate-carbonate deposition in kidney. In this research we studied effect of microgravity on multiplication and calcification of NB in vitro. METHODS: We examined NB cultures in High Aspect Rotating Vessels (HARVs) designed at the NASA's Johnson Space Center, which are designed to stimulate some aspects of microgravity. Multiplication rate and calcium phosphate composition of those NB were compared with NB cultured on stationary and shaker flasks. Collected aliquots of the cultures from different incubation periods were analyzed using spectrophotometer, SEM, TEM, EDX, and x-ray diffraction techniques. RESULTS: The results showed that NB multiplied 4.6x faster in HARVs compared to stationary cultures, and 3.2x faster than shaker flask conditions. X-ray diffraction and EDX analysis showed that the degree of apatite crystal formation and the properties of the apatite depend on the specific culture conditions used. CONCLUSION: We now report an increased multiplication rate of NB in microgravity-simulated conditions. Thus, NB infection may have a potential role in kidney stone formation in crew members during space flights. For further proof to this hypothesis, screening of the NB antigen and antibody level in flight crew before and after flight would be necessary.

Weightlessness↗

The salutary effect of dietary calcium on bone mass in a rat model of simulated weightlessness

Whether supplementation of dietary calcium reduces the differences in bone mass of unweighed limbs and normally weighted limbs, and whether parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D (1,25(OH)2D) respond differently to dietary calcium in unweighted animals in comparison with pair-fed controls was studied. The hind limbs of rats were unweighted by a tail suspension method and diets containing 0.1% to 2.4% calcium. After 2 weeks serum calcium, phosphorus, PTH and 1,25(OH)2D intestinal calcium transport were determined and bone mass, ash weight, and calcium in the tibia, L-1 vertebra, and humerus were measured. No significant differences in body weights were observed among the various groups. Suspended rats maintained constant levels of serum calcium and phosphate over the wide range of dietary calcium. Serum PTH and 1,25(OH)2D and intestinal calcium transport fell as dietary calcium was increased. Bone calcium in the tibia and vertebra from suspended rats remained less than that from pair-fed control. It is suggested that although no striking difference between suspended and control animals was observed in response to dieteary calcium, increasing dietary calcium may reduce the negative impact of unloading on the calcium content of the unweighted bones. The salutary effect of high dietary calcium appears to be due to inhibition of bone resorption rather than to stimulation of bone formation.

Bikle, D. D.↗