Changes in expression of genes involved in apoptosis in activated human T-cells in response to modeled microgravity

Space flights result in remarkable effects on various physiological systems, including a decline in cellular immune functions. Previous studies have shown that exposure to microgravity, both true and modeled, can cause significant changes in numerous lymphocyte functions. The purpose of this study was to search for microgravity-sensitive genes, and specifically for apoptotic genes influenced by the microgravity environment and other genes related to immune response. The experiments were performed on anti-CD3 and IL-2 activated human T cells. To model microgravity conditions we have utilized the NASA rotating wall vessel bioreactor. Control lymphocytes were cultured in static 1g conditions. To assess gene expression we used DNA microarray chip technology. We had shown that multiple genes (approximately 3–8% of tested genes) respond to microgravity conditions by 1.5 and more fold change in expression. There is a significant variability in the response. However, a certain reproducible pattern in gene response could be identified. Among the genes showing reproducible changes in expression in modeled microgravity, several genes involved in apoptosis as well as in immune response were identified. These are IL-7 receptor, Granzyme B, Beta-3-endonexin, Apo2 ligand and STAT1. Possible functional consequences of these changes are discussed.     

Facilities for Simulation of Microgravity in the ESA Ground-Based Facility Programme

Knowledge of the role of gravity in fundamental biological processes and, consequently, the impact of exposure to microgravity conditions provide insight into the basics of the development of life as well as enabling long-term space exploration missions. However, experimentation in real microgravity is expensive and scarcely available; thus, a variety of platforms have been developed to provide, on Earth, an experimental condition comparable to real microgravity. With the aim of simulating microgravity conditions, different ground-based facilities (GBF) have been constructed such as clinostats and random positioning machines as well as magnets for magnetic levitation. Here, we give an overview of ground-based facilities for the simulation of microgravity which were used in the frame of an ESA ground-based research programme dedicated to providing scientists access to these experimental capabilities in order to prepare their space experiments.     

A Sounding Rocket Experiment to Control Boiling by Means of Acoustic Waves

One of the most critical issues when considering long-term space exploration missions is the management and storage of cryogenic propellants. The exposure of storage tanks to radiation and extreme temperatures implies the need of efficient technologies to counteract their effects on the fuel. A potentially dangerous effect for spacecraft operations is the generation of vapor bubbles in cryogenic propellants. We present an experimental setup and procedure to mature a technology based on acoustic waves to control boiling in microgravity.     

Implications of the space radiation environment for human exploration in deep space

Human exploration of the solar system beyond Earth's orbit will entail many risks for the crew on these deep space missions. One of the most significant health risks is exposure to the harsh space radiation environment beyond the protection provided by the Earth's intrinsic magnetic field. Crew on exploration missions will be exposed to a complex mixture of very energetic particles. Chronic exposures to the ever-present background galactic cosmic ray (GCR) spectrum consisting of all naturally occurring chemical elements are combined with sporadic, possibly acute exposures to large fluxes of solar energetic particles, mainly protons and alpha particles. The background GCR environment is mainly a matter of concern for stochastic effects, such as the induction of cancer with subsequent mortality in many cases, and late deterministic effects, such as cataracts and possible damage to the central nervous system. Unfortunately, the actual risks of cancer induction and mortality owing to the very important high-energy heavy ion component of the GCR spectrum are essentially unknown. The sporadic occurrence of extremely large solar energetic particle events (SPE), usually associated with intense solar activity, is also a major concern for the possible manifestation of acute effects from the accompanying high doses of such radiations, especially acute radiation syndrome effects such as nausea, emesis, haemorrhaging or, possibly, even death. In this presentation, an overview of the space radiation environment, estimates of the associated body organ doses and equivalent doses and the potential biological effects on crew in deep space are presented. Possible methods of mitigating these radiations, thereby reducing the associated risks to crew are also described.     

The Impact of Simulated Microgravity on the Growth of Different Genotypes of the Model Legume Plant <Emphasis Type="Italic">Medicago truncatula</Emphasis>

Simulated microgravity has been a useful tool to help understand plant development in altered gravity conditions. Thirty-one genotypes of the legume plant Medicago truncatula were grown in either simulated microgravity on a rotating clinostat, or in a static, vertical environment. Twenty morphological features were measured and compared between these two gravity treatments. Within-species genotypic variation was a significant predictor of the phenotypic response to gravity treatment in 100% of the measured morphological and growth features. In addition, there was a genotype–environment interaction (G × E) for 45% of the response variables, including shoot relative growth rate (p <?0.0005), median number of roots (p ～ 0.02), and root dry mass (p <?0.005). Our studies demonstrate that genotype does play a significant role in M. truncatula morphology and affects the response of plants to the gravity treatment, influencing both the magnitude and direction of the gravity response. These findings are discussed in the context of improving future studies in plant space biology by controlling for genotypic differences. Thus, manipulation of genotype effects, in combination with M. truncatula’s symbiotic relationships with bacteria and fungi, will be important for optimizing legumes for cultivation on long-term space missions.     

Simulation of Microgravity by Magnetic Levitation and Random Positioning: Effect on Human A431 Cell Morphology

Simulation of weightlessness is a desired replenishment for research in microgravity since access to space flights is limited. In real microgravity conditions, the human epidermoid cell line A431 exhibits specific changes in the actin cytoskeleton resulting ultimately in the rounding-up of cells. This rounding of A431 cells was studied in detail during exposure to Random Positioning Machine (RPM) rotation and magnetic levitation. Random rotation and magnetic levitation induced similar changes in the actin morphology of A431 cells that were also described in real microgravity. A transient process of cell rounding and renewed spreading was observed in time, illustrated by a changing actin cytoskeleton and variation in the presence of focal adhesions. However, side effects of both methods easily can lead to false linking of cellular responses to simulated microgravity. Therefore further characterization of both methods is required.     

Real-Time Video-Microscopy of Migrating Immune Cells in Altered Gravity During Parabolic Flights

In our project we developed a technical equipment which allows to visualize migration of cells in real-time video-microscopy during altered gravity conditions of NOVESPACE Airbus A300 ZERO-G parabolic flights. For validation of the experimental device we have used fast moving human neutrophils as example, because their migration is fundamental to keep the organism under immunological surveillance. Their migration is indispensable for immune effector function, where the cells leave the blood vessels and navigate to places of infection to fulfill their main task of phagocytosis. Thereby, we have analyzed if their migration is affected during altered gravity conditions and if pharmacological modification of cytoskeletal dynamics influences neutrophil migratory activity. Whereas we detected no change in neutrophil locomotory behaviour in microgravity, we found a significant inhibitory influence of hypergravity, irrespective of the chemical stimulus used. Our results suggest that hypergravity, following a microgravity environment, could represent a hazard to the human immune system function. Thus, our cell migration assay offers an optimum experimental device for studying the migratory activity and underlying signal transduction mechanisms of neutrophils to assess the immunological fitness of humans in space to fight infection, but also for investigating the locomotion of other cell types or unicellular organisms such as ciliates.     

The “ageing” experiment in the spanish soyuz mission to the international space station

Human exploration of outer space will eventually take place. In preparation for this endeavour, it is important to establish the nature of the biological response to a prolonged exposure to the space environment. In one of the recent Soyuz Missions to serve the International Space Station (ISS), the Spanish Soyuz mission in October 2003, we exposed four groups of Drosophila male imagoes to microgravity during the almost eleven days of the Cervantes mission to study their motility behaviour. The groups were three of young flies and one of mature flies, In previous space experiments, we have shown that when imagoes are exposed to microgravity they markedly change their behaviour by increasing their motility, especially if subjected to these conditions immediately after hatching. The constraints of the current Soyuz flights made it impossible to study the early posthatching period. A low temperature cold transport was incorporated as a possible way out of this constraint. It turned out that on top of the space flight effects, the cold treatment by itself, modifies the motility behaviour of the flies. Although the four groups increased their motility, the young flies did it in a much lower extent than the mature flies that had not been exposed to the low temperature during transportation. Nevertheless, the flies flown in the ISS are still more active than the parallel ground controls. As a consequence of the lower motility stimulation in this experiment, a likely consequence of the cold transport step, no effects on the life spans of the flown flies were detected. Together with previous results, this study confirms that high levels of motility behaviour are necessary to produce significant decreases in fly longevity.     

ARADISH - Development of a Standardized Plant Growth Chamber for Experiments in Gravitational Biology Using Ground Based Facilities

Plant development strongly relies on environmental conditions. Growth of plants in Biological Life Support Systems (BLSS), which are a necessity to allow human survival during long-term space exploration missions, poses a particular problem for plant growth, as in addition to the traditional environmental factors, microgravity (or reduced gravity such as on Moon or Mars) and limited gas exchange hamper plant growth. Studying the effects of reduced gravity on plants requires real or simulated microgravity experiments under highly standardized conditions, in order to avoid the influence of other environmental factors. Analysis of a large number of biological replicates, which is necessary for the detection of subtle phenotypical differences, can so far only be achieved in Ground Based Facilities (GBF). Besides different experimental conditions, the usage of a variety of different plant growth chambers was a major factor that led to a lack of reproducibility and comparability in previous studies. We have developed a flexible and customizable plant growth chamber, called ARAbidopsis DISH (ARADISH), which allows plant growth from seed to seedling, being realized in a hydroponic system or on Agar. By developing a special holder, the ARADISH can be used for experiments with Arabidopsis thaliana or a plant with a similar habitus on common GBF hardware, including 2D clinostats and Random Positioning Machines (RPM). The ARADISH growth chamber has a controlled illumination system of red and blue light emitting diodes (LED), which allows the user to apply defined light conditions. As a proof of concept we tested a prototype in a proteomic experiment in which plants were exposed to simulated microgravity or a 90° stimulus. We optimized the design and performed viability tests after several days of growth in the hardware that underline the utility of ARADISH in microgravity research.     

Flowering in space

The reproductive success of plants is often dependent on their flowering time being adapted to the terrestrial environment, in which gravity remain constant. Whether plants can follow the same rule to determine their flowering time under microgravity in space is unknown. Although numerous attempts have been made to grow a plant through a complete life cycle in space, apparently no published information exists concerning the flowering control of plants under microgravity in space. Here, we focused on two aspects. Firstly the environmental and intrinsic factors under microgravity related to flowering control. Secondly, the plant-derived regulators are involved in flowering control under microgravity condition. The potential environmental and intrinsic factors affect plant flowering under microgravity may include light, biological circadian clock as well as long-distance signaling, while the plant-derived flowering regulators in response to microgravity could include gibberellic acid, ethylene, microRNA and sugar. The results we have obtained from the space experiments on board the Chinese recoverable satellites (the SJ-8 and the SJ-10) and the experiment on the Chinese space lab TG-2 are also introduced. We conclude by suggesting that long-term space experiments from successive generations and a systematic analysis of regulatory networks at the molecular level is needed to understand the mechanism of plant flowering control under microgravity conditions in space.     

Approach and issues relating to shield material design to protect astronauts from space radiation

One major obstacle to human space exploration is the possible limitations imposed by the adverse effects of long-term exposure to the space environment. Even before human spaceflight began, the potentially brief exposure of astronauts to the very intense random solar energetic particle (SEP) events was of great concern. A new challenge appears in deep space exploration from exposure to the low-intensity heavy-ion flux of the galactic cosmic rays (GCR) since the missions are of long duration and the accumulated exposures can be high. Since aluminum (traditionally used in spacecraft to avoid potential radiation risks) leads to prohibitively expensive mission launch costs, alternative materials need to be explored. An overview of the materials related issues and their impact on human space exploration will be given.     

From hypergravity to microgravity: Choosing the suitable simulator

Cardiovascular system functions are impaired in altered gravity conditions. In particular endothelial cells play a major role being responsible for the integrity of the vascular wall. Due to obvious difficulties in performing continuous and exhaustive experiments in space, most of the available data have been obtained so far using various simulators of hypergravity and microgravity (µg) conditions. The consistency of the data resides on the reliability of the simulator, being a critical point in the development of the research. We exposed the cell cultures to 1) hypergravity (launch condition) using MidiCAR at Dutch Experiment Support Center (DESC, NL); 2) simulated µg using the Rotating Wall Vessel (RWV) and the Random Positioning Machine (RPM). We used two different cellular models: human umbilical vein endothelial cells (HUVEC) and human leukocytes (U937). Only few experiments on cells using RPM have been reported. To assess the RPM best operative parameters we considered data from experiments in space on U937 as reference standard. Differently, cultures in modelled µg using RWV have been extensively reported. Our data on HUVEC indicate that the two µg simulators provide analogous results in terms of proliferation and cytoskeletal organization. Finally, to investigate the effects of spaceflight on different human cells, we developed a spaceflight-like protocol consisting of an initial hypergravity phase (launch), followed by a µg simulation (orbital flight). Using this protocol, results show that hypergravity limits in our models the effects on proliferation induced by modelled µg.     

Drop Tower Experiment to Study the Capillary Flow in Symmetrical and Asymmetrical Channels: Experimental Set-up and Preliminary Results

Owing to the development of the space exploration activities, the in-orbit management of fluids such as the transportation of propellant liquid in microgravity becomes the important direction of microgravity fluid research, and one of main problems is the stability behaviors of free surface flow in capillary channel of PMD. In the present study, an experiment set-up of the fluid transport with two different capillary channels has been developed on the Beijing Drop Tower platform. Both symmetrical and asymmetrical flow channels, with the same cross-sectional areas and lengths and different cross-sectional geometries were used and HFE-7500 is chosen as test liquid. 10 times of the drop-down experiments were performed for investigation of the capillary flow characters in different volumetric flow rates, and the three main patterns of capillary flows: subcritical flow, critical flow and supercritical flow were found in experiments, these patterns are distinguished by the movement of the point of lowest surface over time. Meanwhile, the critical flow rates at which free surface becomes instable observed in our experiments are (1) 2.7 ±0.2ml/s for the critical flow rate of asymmetrical channel; and (2) 2.2 ±0.2ml/s for symmetrical channel flow, respectively.     

Nanoparticle-induced exosomes target antigen-presenting cells to initiate Th1-type immune activation

The mechanisms associated with the induction of systemic immune responses by nanoparticles are not fully understood, but their elucidation is critical to address safety issues associated with the broader medical application of nanotechnology. In this study, a key role of nanoparticle-induced exosomes (extracellularly secreted membrane vesicles) as signaling mediators in the induction of T helper cell type 1 (Th1) immune activation is demonstrated. In vivo exposure to magnetic iron oxide nanoparticles (MIONs) results in significant exosome generation in the alveolar region of Balb/c mice. These act as a source of nanoparticle-induced, membrane-bound antigen/signaling cargo, which transfer their components to antigen-presenting cells (APCs) in the reticuloendothelial system. Through exosome-initiated signals, immature dendritic cells (iDCs) undergo maturation and differentiation to the DC1 subtype, while macrophages go through classical activation and differentiation to the M1 subtype. Simultaneously, iDCs and macrophages release various Th1 cytokines (including interleukin-12 and tumor necrosis factor α) driving T-cell activation and differentiation. Activated APCs (especially DC1 and M1 subtypes) consequently prime T-cell differentiation towards a Th1 subtype, thereby resulting in an orchestrated Th1-type immune response. Th1-polarized immune activation is associated with delayed-type hypersensitivity, which might underlie the long-term inflammatory effects frequently associated with nanoparticle exposure. These studies suggest that nanoparticle-induced exosomes provoke the immune activation and inflammatory responses that can accompany nanoparticle exposure.     

Radiolabeling,quality control,and biodistribution of <Superscript>99m</Superscript>Tc-sulfadiazine as an infection imaging agent

Abstract—Sulfadiazine (antibiotic used for treating bacterial infections) was labeled with ^99mTc with the aim of the development of a new radiopharmaceutical for infection imaging. The influence of the reaction parameters such as the substrate and SnCl₂·2H₂O concentrations, pH of the reaction mixture, and reaction time on the labeling yield was examined, and the labeling conditions were optimized. The maximum radiochemical yield of ^99mTc-sulfadiazine (94.7%) was obtained by using 50 µg of SnCl₂·2H₂O and 1 mg of sulfadiazine at pH 5. The radiochemical purity of the labeled compound was evaluated by ITLC and HPLC. The biological distribution of ^99mTc-sulfadiazine in mice with experimentally induced Staphylococcus aureus infection in the right thigh was studied, and the bacterial infected thigh/normal thigh (target-to-nontarget, T/NT) ratio was evaluated. The T/NT ratio for ^99mTc-sulfadiazine was found to be 3.6, which is comparable to the commercially available ^99mTc-ciprofloxacin (3.8), indicating that ^99mTc-sulfadiazine can be used for infection imaging.     

The Lymphocyte Story – an Overview of Selected Highlights on the in Vitro Activation of Human Lymphocytes in Space

Since the first flight of humans into space it is known that space flight affects the immune system; especially a weakening of the reactivity of T-lymphocytes after flight has been observed. In an in vitro experiment, proposed by Augusto Cogoli and flown in Spacelab-1 in 1983, the activation of T-lymphocytes was found to be strongly inhibited in microgravity. This surprising result triggered extended investigations in space and on the ground by us and other research teams. T-cells are that subpopulation of lymphocytes responsible for the activation of the specific immune system. The mechanism of T-cell activation is very complex; 3 different signals are required as well as an interaction between T-lymphocytes and monocytes. Cell motility based on a continuous rearrangement of the cytoskeletal network within the cell is essential for cell-cell contacts. The objective of all our experiments performed on different platforms in space as well as in simulated microgravity on ground was to understand and explain the dysfunction of the cell activation under reduced gravity conditions. On sounding rockets we have studied the influence of microgravity on the delivery of the first signal, the motility of lymphocytes as well as changes in the cytoskeletal structure and early gene expression. On long term missions we investigated many aspects of the delivery of the 2 ^nd and 3 ^rd signal, including motility and aggregate formation of lymphocytes, interaction of lymphocytes with monocytes, motility of monocytes and changes in different cytoskeletal structures.     

Amperometric biosensor based on carbon nanotubes coated with polyaniline/dendrimer-encapsulated Pt nanoparticles for glucose detection

A novel amperometric glucose biosensor based on the nanocomposites of multi-wall carbon nanotubes (CNT) coated with polyaniline (PANI) and dendrimer-encapsulated Pt nanoparticles (Pt-DENs) is prepared. CNT coated with protonated PANI is in situ synthesized and Pt-DENs is absorbed on PANI/CNT composite surface by self-assembly method. Then Glucose oxidase (GOx) is crosslink-immobilizated onto Pt-DENs/PANI/CNT composite film. The results show that the fabricated GOx/Pt-DENs/PANI/CNT electrode exhibits excellent response performance to glucose, such as low detection limit (0.5 µM), wide linear range (1 µM–12 mM), short response time (about 5 s), high sensitivity (42.0 µA mM^? 1 cm^? 2) and stability (83% remains after 3 weeks).     

Radiation dosimetry for microbial experiments in the International Space Station using different etched track and luminescent detectors

The laboratory of Microbiology at SCK.CEN, in collaboration with different universities, participates in several ESA programmes with bacterial experiments that are carried out in the International Space Station (ISS). The main objective of these programmes is to study the effects of space flight conditions such as microgravity and cosmic radiation on the general behaviour of model bacteria. To measure the radiation doses received by the bacteria, different detectors accompanied the microbiological experiments. The results obtained during two space flight missions are discussed. This dosimetry experiment was a collaboration between different institutes so that the doses could be estimated by different techniques. For measurement of the high linear energy transfer (LET) doses (>10 keV microm(-1)), two types of etched track detectors were used. The low LET part of the spectrum was measured by three types of thermoluminescent detectors ((7)LiF:Mg,Ti; (7)LiF:Mg,Cu,P; Al(2)O(3):C) and by the optically stimulated luminescence technique using Al(2)O(3):C detectors.