The enigmatic Placozoa part 2: Exploring evolutionary controversies and promising questions on earth and in space.

The placozoan Trichoplax adhaerens has been bridging gaps between research disciplines like no other animal. As outlined in part 1, placozoans have been subject of hot evolutionary debates and placozoans have challenged some fundamental evolutionary concepts. Here in part 2 we discuss the exceptional genetics of the phylum Placozoa and point out some challenging model system applications for the best known species, Trichoplax adhaerens.

The enigmatic Placozoa part 1: Exploring evolutionary controversies and poor ecological knowledge.

The placozoan Trichoplax adhaerens is a tiny hairy plate and more simply organized than any other living metazoan. After its original description by F.E. Schulze in 1883, it attracted attention as a potential model for the ancestral state of metazoan organization, the "Urmetazoon". Trichoplax lacks any kind of symmetry, organs, nerve cells, muscle cells, basal lamina, and extracellular matrix. Furthermore, the placozoan genome is the smallest (not secondarily reduced) genome of all metazoan genomes. It harbors a remarkably rich diversity of genes and has been considered the best living surrogate for a metazoan ancestor genome. The phylum Placozoa presently harbors three formally described species, while several dozen "cryptic" species are yet awaiting their description. The phylogenetic position of placozoans has recently become a contested arena for modern phylogenetic analyses and view-driven claims. Trichoplax offers unique prospects for understanding the minimal requirements of metazoan animal organization and their corresponding malfunctions.

Simulating Space Conditions Evokes Different DNA Damage Responses in Immature and Mature Cells of the Human Hematopoietic System.

The impact of space radiation and microgravity on DNA damage responses has been discussed controversially, largely due to the variety of model systems engaged. Here, we performed side-by-side analyses of human hematopoietic stem/progenitor cells (HSPC) and peripheral blood lymphocytes (PBL) cultivated in a 2D clinostat to simulate microgravity before, during and after photon and particle irradiation. We demonstrate that simulated microgravity (SMG) accelerates the early phase of non-homologous end joining (NHEJ)-mediated repair of simple, X-ray-induced DNA double-strand breaks (DSBs) in PBL, while repair kinetics in HSPC remained unaltered. Repair acceleration was lost with increasing LET of ion exposures, which increases the complexity of DSBs, precluding NHEJ and requiring end resection for successful repair. Such cell-type specific effect of SMG on DSB repair was dependent on the NF-кB pathway pre-activated in PBL but not HSPC. Already under unperturbed growth conditions HSPC and PBL suffered from SMG-induced replication stress associated with accumulation of single-stranded DNA and DSBs, respectively. We conclude that in PBL, SMG-induced DSBs promote repair of radiation-induced damage in an adaptive-like response. HSPC feature SMG-induced single-stranded DNA and FANCD2 foci, i.e., markers of persistent replication stress and senescence that may contribute to a premature decline of the immune system in space.

Hardware Development for Plant Cultivation Allowing In Situ Fluorescence Analysis of Calcium Fluxes in Plant Roots Under Microgravity and Ground-Control Conditions.

Maintaining an optimal leaf and stem orientation to yield a maximum photosynthetic output is accomplished by terrestrial plants using sophisticated mechanisms to balance their orientation relative to the Earth's gravity vector and the direction of sunlight. Knowledge of the signal transduction chains of both gravity and light perception and how they influence each other is essential for understanding plant development on Earth and plant cultivation in space environments. However, in situ analyses of cellular signal transduction processes in weightlessness, such as live cell imaging of signaling molecules using confocal fluorescence microscopy, require an adapted experimental setup that meets the special requirements of a microgravity environment. In addition, investigations under prolonged microgravity conditions require extensive resources, are rarely accessible, and do not allow for immediate sample preparation for the actual microscopic analysis. Therefore, supply concepts are needed that ensure both the viability of the contained plants over a longer period of time and an unhindered microscopic analysis in microgravity. Here, we present a customized supply unit specifically designed to study gravity-induced Ca2+ mobilization in roots of Arabidopsis thaliana. The unit can be employed for ground-based experiments, in parabolic flights, on sounding rockets, and probably also aboard the International Space Station.

The impact of altered gravity and vibration on endothelial cells during a parabolic flight.

BACKGROUND: Endothelial cells (EC) cultured under altered gravity conditions show a cytoskeletal disorganization and differential gene expression (short-term effects), as well as apoptosis in adherently growing EC or formation of tubular 3D structures (long-term effects). METHODS: Investigating short-term effects of real microgravity, we exposed EC to parabolic flight maneuvers and analysed them on both protein and transcriptional level. The effects of hypergravity and vibration were studied separately. RESULTS: Pan-actin and tubulin proteins were elevated by vibration and down-regulated by hypergravity. ß-Actin was reduced by vibration. Moesin protein was reduced by both vibration and hypergravity, ezrin potein was strongly elevated under vibration. Gene expression of ACTB, CCND1, CDC6, CDKN1A, VEGFA, FLK-1, EZR, ITBG1, OPN, CASP3, CASP8, ANXA2, and BIRC5 was reduced under vibration. With the exception of CCNA2, CCND1, MSN, RDX, OPN, BIRC5, and ACTB all investigated genes were downregulated by hypergravity. After one parabola (P) CCNA2, CCND1, CDC6, CDKN1A, EZR, MSN, OPN, VEGFA, CASP3, CASP8, ANXA1, ANXA2, and BIRC5 were up-, while FLK1 was downregulated. EZR, MSN, OPN, ANXA2, and BIRC5 were upregulated after 31P. CONCLUSIONS: Genes of the cytoskeleton, angiogenesis, extracellular matrix, apoptosis, and cell cycle regulation were affected by parabolic flight maneuvers. We show that the microgravity stimulus is stronger than hypergravity/vibration.

Short-term weightlessness produced by parabolic flight maneuvers altered gene expression patterns in human endothelial cells.

This study focused on the effects of short-term microgravity (22 s) on the gene expression and morphology of endothelial cells (ECs) and evaluated gravisensitive signaling elements. ECs were investigated during four German Space Agency (Deutsches Zentrum für Luft- und Raumfahrt) parabolic flight campaigns. Hoechst 33342 and acridine orange/ethidium bromide staining showed no signs of cell death in ECs after 31 parabolas (P31). Gene array analysis revealed 320 significantly regulated genes after the first parabola (P1) and P31. COL4A5, COL8A1, ITGA6, ITGA10, and ITGB3 mRNAs were down-regulated after P1. EDN1 and TNFRSF12A mRNAs were up-regulated. ADAM19, CARD8, CD40, GSN, PRKCA (all down-regulated after P1), and PRKAA1 (AMPKα1) mRNAs (up-regulated) provide a very early protective mechanism of cell survival induced by 22 s microgravity. The ABL2 gene was significantly up-regulated after P1 and P31, TUBB was slightly induced, but ACTA2 and VIM mRNAs were not changed. ß-Tubulin immunofluorescence revealed a cytoplasmic rearrangement. Vibration had no effect. Hypergravity reduced CARD8, NOS3, VASH1, SERPINH1 (all P1), CAV2, ADAM19, TNFRSF12A, CD40, and ITGA6 (P31) mRNAs. These data suggest that microgravity alters the gene expression patterns and the cytoskeleton of ECs very early. Several gravisensitive signaling elements, such as AMPKα1 and integrins, are involved in the reaction of ECs to altered gravity.

Human neural network activity reacts to gravity changes in vitro.

During spaceflight, humans experience a variety of physiological changes due to deviations from familiar earth conditions. Specifically, the lack of gravity is responsible for many effects observed in returning astronauts. These impairments can include structural as well as functional changes of the brain and a decline in cognitive performance. However, the underlying physiological mechanisms remain elusive. Alterations in neuronal activity play a central role in mental disorders and altered neuronal transmission may also lead to diminished human performance in space. Thus, understanding the influence of altered gravity at the cellular and network level is of high importance. Previous electrophysiological experiments using patch clamp techniques and calcium indicators have shown that neuronal activity is influenced by altered gravity. By using multi-electrode array (MEA) technology, we advanced the electrophysiological investigation covering single-cell to network level responses during exposure to decreased (micro-) or increased (hyper-) gravity conditions. We continuously recorded in real-time the spontaneous activity of human induced pluripotent stem cell (hiPSC)-derived neural networks in vitro. The MEA device was integrated into a custom-built environmental chamber to expose the system with neuronal cultures to up to 6 g of hypergravity on the Short-Arm Human Centrifuge at the DLR Cologne, Germany. The flexibility of the experimental hardware set-up facilitated additional MEA electrophysiology experiments under 4.7 s of high-quality microgravity (10-6 to 10-5 g) in the Bremen drop tower, Germany. Hypergravity led to significant changes in activity. During the microgravity phase, the mean action potential frequency across the neural networks was significantly enhanced, whereas different subgroups of neurons showed distinct behaviors, such as increased or decreased firing activity. Our data clearly demonstrate that gravity as an environmental stimulus triggers changes in neuronal activity. Neuronal networks especially reacted to acute changes in mechanical loading (hypergravity) or de-loading (microgravity). The current study clearly shows the gravity-dependent response of neuronal networks endorsing the importance of further investigations of neuronal activity and its adaptive responses to micro- and hypergravity. Our approach provided the basis for the identification of responsible mechanisms and the development of countermeasures with potential implications on manned space missions.

Differential gene regulation under altered gravity conditions in follicular thyroid cancer cells: relationship between the extracellular matrix and the cytoskeleton.

Extracellular matrix proteins, adhesion molecules, and cytoskeletal proteins form a dynamic network interacting with signalling molecules as an adaptive response to altered gravity. An important issue is the exact differentiation between real microgravity responses of the cells or cellular reactions to hypergravity and/or vibrations. To determine the effects of real microgravity on human cells, we used four DLR parabolic flight campaigns and focused on the effects of short-term microgravity (22 s), hypergravity (1.8 g), and vibrations on ML-1 thyroid cancer cells. No signs of apoptosis or necrosis were detectable. Gene array analysis revealed 2,430 significantly changed transcripts. After 22 s microgravity, the F-actin and cytokeratin cytoskeleton was altered, and ACTB and KRT80 mRNAs were significantly upregulated after the first and thirty-first parabolas. The COL4A5 mRNA was downregulated under microgravity, whereas OPN and FN were significantly upregulated. Hypergravity and vibrations did not change ACTB, KRT-80 or COL4A5 mRNA. MTSS1 and LIMA1 mRNAs were downregulated/slightly upregulated under microgravity, upregulated in hypergravity and unchanged by vibrations. These data indicate that the graviresponse of ML-1 cells occurred very early, within the first few seconds. Downregulated MTSS1 and upregulated LIMA1 may be an adaptive mechanism of human cells for stabilizing the cytoskeleton under microgravity conditions.

The MAPHEUS module CellFix for studying the influence of altered gravity on the physiology of single cells.

Gravity is the only constant stimulus during the evolution of life. To investigate the impact of the absence of gravity on living systems, their molecular and morphological status has to be studied under microgravity conditions. The experiment unit CellFix was developed in order to provide the possibility of exposure and chemical fixation of small biological systems, such as neurons, stem cells, small animals, yeast cultures, plants, etc., at dedicated time points during a sounding rocket flight. The current version of CellFix consists of two culture bags containing cell cultures in a temperature-controlled pressure vessel. The biosystems in the culture bags can be fixed by pumping the fixative [e.g., paraformaldehyde (PFA), methanol, RNAlater, or others] from a connected bag into the cell suspension. The mechatronic basis of the experiment unit is constructed from compartments of the shelf parts. Open source microcontroller systems (Arduino) or gear pumps, accumulators, etc., from the model making sector are affordable and reliable components to build up an experiment on an unmanned space mission such as a sounding rocket flight. Also, new technologies such as fused deposition modeling were used to construct structures and brackets, which were tested successfully in environmental tests and real space flights (MAPHEUS 7 and 8 sounding rocket missions). In combination with the possibility to handle the experiment as a late access insert in a standardized rocket compartment, CellFix provides a multiusable experiment unit for performing life science experiments in space.

ARABIDOMICS-A new experimental platform for molecular analyses of plants in drop towers, on parabolic flights, and sounding rockets.

Plants represent an essential part of future life support systems that will enable human space travel to distant planets and their colonization. Therefore, insights into changes and adaptations of plants in microgravity are of great importance. Despite considerable efforts, we still know very little about how plants respond to microgravity environments on the molecular level, partly due to a lack of sufficient hardware and flight opportunities. The plant Arabidopsis thaliana, the subject of this study, represents a well-studied model organism in gravitational biology, particularly for the analysis of transcriptional and metabolic changes. To overcome the limitations of previous plant hardware that often led to secondary effects and to allow for the extraction not only of RNA but also of phytohormones and proteins, we developed a new experimental platform, called ARABIDOMICS, for exposure and fixation under altered gravity conditions. Arabidopsis seedlings were exposed to hypergravity during launch and microgravity during the free-fall period of the MAPHEUS 5 sounding rocket. Seedlings were chemically fixed inflight at defined time points, and RNA and phytohormones were subsequently analyzed in the laboratory. RNA and phytohormones extracted from the fixed biological samples were of excellent quality. Changes in the phytohormone content of jasmonate, auxin, and several cytokinins were observed in response to hypergravity and microgravity.

The gravity dependence of pharmacodynamics: the integration of lidocaine into membranes in microgravity.

To realize long-term manned space missions, e.g. to Mars, some important questions about pharmacology under conditions of different gravity will have to be answered to ensure safe usage of pharmaceuticals. Experiments on the International Space Station showed that the pharmacokinetics of drugs are changed in microgravity. On Earth, it is well known that the incorporation of substances into cellular membranes depends on membrane fluidity, therefore the finding that membrane fluidity is gravity dependent possibly has effects on pharmacodynamics of hydrophobic and amphiphilic substances in microgravity. To validate a possible effect of gravity on pharmacodynamics, experiments have been carried out to investigate the incorporation of lidocaine into plain lipid membranes under microgravity conditions. In microgravity, the induced increase in membrane fluidity associated with lidocaine incorporation is smaller compared to 1g controls. This experiment concerning the gravity dependence of pharmacodynamics in real microgravity clearly shows that the incorporation of amphipathic drugs into membranes is changed in microgravity. This might have significant impact on the pharmacology of drugs during long-term space missions and has to be investigated in more detail to be able to assess possible risks.

The influence of nitrogen concentration and precipitation on fertilizer production from urine using a trickling filter.

Planetary habitation requires technology to maintain natural microbial processes, which make nutrients from biowaste available for plant cultivation. This study describes a 646 day experiment, in which trickling filters were monitored for their ability to mineralize nitrogen when loaded with artificial urine solutions of different concentrations (40, 60, 80 and 100% v/v). Former studies have indicated that increasing urine concentrations slow nitrogen conversion rates and induce growing instability. In the current experiment, nitrogen conversion rates, measured as nitrate production/day, did not differ between concentration levels and increasing instability was not observed. Instead, the buffering capacity of the mussel shells added as buffer system (∼75% calcium carbonate) increased with increasing concentrations of synthetic urine possibly due to the higher phosphate content. The intensified precipitation of calcium phosphates seems to promote carbonate dissolution leading to improved buffering. For space applications, the precipitation of calcium phosphates is not desirable as for the phosphate to be available to the plants the precipitate must be treated with hazardous substances. With regard to terrestrial agriculture the process-integrated phosphate precipitation is a possibility to separate the macronutrients nitrogen and phosphate without addition of other chemicals. Thus, the described process offers a simple and cost-effective approach to fertilizer production from biogenic residues like slurry.

Pyrocystis noctiluca represents an excellent bioassay for shear forces induced in ground-based microgravity simulators (clinostat and random positioning machine).

Ground-based facilities, such as clinostats and random positioning machines aiming at simulating microgravity conditions, are tools to prepare space experiments and identify gravity-related signaling pathways. A prerequisite is that the facilities are operated in an appropriate manner and potentially induced non-gravitational effects, such as shearing forces, have to be taken into account. Dinoflagellates, here P. noctiluca, as fast and sensitive reporter system for shear stress and hydrodynamic gradients, were exposed on a clinostat (constant rotation around one axis, 60 rpm) or in a random positioning machine, that means rotating around two axes, whose velocity and direction were chosen at random. Deformation of the cell membrane of P. noctiluca due to shear stress results in a detectable bioluminescence emission. Our results show that the amount of mechanical stress is higher on an random positioning machine than during constant clinorotation, as revealed by the differences in photon counts. We conclude that one axis clinorotation induced negligible non-gravitational effects in the form of shear forces in contrast to random operation modes tested. For the first time, we clearly visualized the device-dependent occurrence of shear forces by means of a bioassay, which have to be considered during the definition of an appropriate simulation approach and to avoid misinterpretation of results.

Long term stability of Oligo (dT) 25 magnetic beads for the expression analysis of Euglena gracilis for long term space projects.

The unicellular freshwater flagellate Euglena gracilis has a highly developed sensory system. The cells use different stimuli such as light and gravity to orient themselves in the surrounding medium to find areas for optimal growth. Due to the ability to produce oxygen and consume carbon dioxide, Euglena is a suitable candidate for life support systems. Participation in a long-term space experiment would allow for the analysis of changes and adaptations to the new environment, and this could bring new insights into the mechanism of perception of gravity and the associated signal transduction chain. For a molecular analysis of transcription patterns, an automated system is necessary, capable of performing all steps from taking a sample, processing it and generating data. One of the developmental steps is to find long-term stable reagents and materials and test them for stability at higher-than-recommended temperature conditions during extended storage time. We investigated the usability of magnetic beads in an Euglena specific lysis buffer after addition of the RNA stabilizer Dithiothreitol over 360 days and the lysis buffer with the stabilizer alone over 455 days at the expected storage temperature of 19 °C. We can claim that the stability is not impaired at all after an incubation period of over one year. This might be an interesting result for researchers who have to work under non-standard lab conditions, as in biological or medicinal fieldwork.

Natural microbial populations in a water-based biowaste management system for space life support.

The reutilization of wastewater is a key issue with regard to long-term space missions and planetary habitation. This study reports the design, test runs and microbiological analyses of a fixed bed biofiltration system which applies pumice grain (16-25 mm grain size, 90 m(2)/m(3) active surface) as matrix and calcium carbonate as buffer. For activation, the pumice was inoculated with garden soil known to contain a diverse community of microorganisms, thus enabling the filtration system to potentially degrade all kinds of organic matter. Current experiments over 194 days with diluted synthetic urine (7% and 20%) showed that the 7% filter units produced nitrate slowly but steadily (max. 2191 mg NO3-N/day). In the 20% units nitrate production was slower and less stable (max. 1411 mg NO3-N/day). 84% and 76% of the contained nitrogen was converted into nitrate. The low conversion rate is assumed to be due to the high flow rate, which keeps the biofilm on the pumice thin. At the same time the thin biofilm seems to prevent the activity of denitrifiers implicating the existence of a trade off between rate and the amount of nitrogen loss. Microbiological analyses identified a comparatively low number of species (26 in the filter material, 12 in the filtrate) indicating that urine serves as a strongly selective medium and filter units for the degradation of mixed feedstock have to be pre-conditioned on the intended substrates from the beginning.

Common Effects on Cancer Cells Exerted by a Random Positioning Machine and a 2D Clinostat.

In this study we focused on gravity-sensitive proteins of two human thyroid cancer cell lines (ML-1; RO82-W-1), which were exposed to a 2D clinostat (CLINO), a random positioning machine (RPM) and to normal 1g-conditions. After a three (3d)- or seven-day-culture (7d) on the two devices, we found both cell types growing three-dimensionally within multicellular spheroids (MCS) and also cells remaining adherent (AD) to the culture flask, while 1g-control cultures only formed adherent monolayers, unless the bottom of the culture dish was covered by agarose. In this case, the cytokines IL-6 and IL-8 facilitated the formation of MCS in both cell lines using the liquid-overlay technique at 1g. ML-1 cells grown on the RPM or the CLINO released amounts of IL-6 and MCP-1 into the supernatant, which were significantly elevated as compared to 1g-controls. Release of IL-4, IL-7, IL-8, IL-17, eotaxin-1 and VEGF increased time-dependently, but was not significantly influenced by the gravity conditions. After 3d on the RPM or the CLINO, an accumulation of F-actin around the cellular membrane was detectable in AD cells of both cell lines. IL-6 and IL-8 stimulation of ML-1 cells for 3d and 7d influenced the protein contents of ß1-integrin, talin-1, Ki-67, and beta-actin dose-dependently in adherent cells. The ß1-integrin content was significantly decreased in AD and MCS samples compared with 1g, while talin-1 was higher expressed in MCS than AD populations. The proliferation marker Ki-67 was elevated in AD samples compared with 1g and MCS samples. The ß-actin content of R082-W-1 cells remained unchanged. ML-1 cells exhibited no change in ß-actin in RPM cultures, but a reduction in CLINO samples. Thus, we concluded that simulated microgravity influences the release of cytokines in follicular thyroid cancer cells, and the production of ß1-integrin and talin-1 and predicts an identical effect under real microgravity conditions.

Impact of a high magnetic field on the orientation of gravitactic unicellular organisms--a critical consideration about the application of magnetic fields to mimic functional weightlessness.

The gravity-dependent behavior of Paramecium biaurelia and Euglena gracilis have previously been studied on ground and in real microgravity. To validate whether high magnetic field exposure indeed provides a ground-based facility to mimic functional weightlessness, as has been suggested earlier, both cell types were observed during exposure in a strong homogeneous magnetic field (up to 30 T) and a strong magnetic field gradient. While swimming, Paramecium cells were aligned along the magnetic field lines; orientation of Euglena was perpendicular, demonstrating that the magnetic field determines the orientation and thus prevents the organisms from the random swimming known to occur in real microgravity. Exposing Astasia longa, a flagellate that is closely related to Euglena but lacks chloroplasts and the photoreceptor, as well as the chloroplast-free mutant E. gracilis 1F, to a high magnetic field revealed no reorientation to the perpendicular direction as in the case of wild-type E. gracilis, indicating the existence of an anisotropic structure (chloroplasts) that determines the direction of passive orientation. Immobilized Euglena and Paramecium cells could not be levitated even in the highest available magnetic field gradient as sedimentation persisted with little impact of the field on the sedimentation velocities. We conclude that magnetic fields are not suited as a microgravity simulation for gravitactic unicellular organisms due to the strong effect of the magnetic field itself, which masks the effects known from experiments in real microgravity.

Interleukin-6 expression under gravitational stress due to vibration and hypergravity in follicular thyroid cancer cells.

It is known that exposing cell lines in vitro to parabolic flights changes their gene expression and protein production patterns. Parabolic flights and spaceflight in general are accompanied by transient hypergravity and vibration, which may impact the cells and therefore, have to be considered too. To estimate the possible impact of transient hypergravity and vibration, we investigated the effects of these forces separately using dedicated ground-based facilities. We placed follicular thyroid ML-1 and CGTH W-1 cancer cells in a specific centrifuge (MuSIC Multi Sample Incubator Centrifuge; SAHC Short Arm Human Centrifuge) simulating the hypergravity phases that occur during one (P1) and 31 parabolas (P31) of parabolic flights, respectively. On the Vibraplex device, the same cell lines were treated with vibration waves corresponding to those that occur during a whole parabolic flight lasting for two hours. After the various treatments, cells were harvested and analyzed by quantitative real-time PCR, focusing on the genes involved in forming (ACTB, MYO9, TUBB, VIM, TLN1, and ITGB1) and modulating (EZR, RDX, and MSN) the cytoskeleton, as well as those encoding growth factors (EGF, CTGF, IL6, and IL8) or protein kinases (PRKAA1 and PRKCA). The analysis revealed alterations in several genes in both cell lines; however, fewer genes were affected in ML-1 than CGTH W-1 cells. Interestingly, IL6 was the only gene whose expression was changed in both cell lines by each treatment, while PKCA transcription remained unaffected in all experiments. We conclude that a PKCa-independent mechanism of IL6 gene activation is very sensitive to physical forces in thyroid cells cultured in vitro as monolayers.

Changes in morphology, gene expression and protein content in chondrocytes cultured on a random positioning machine.

Tissue engineering of chondrocytes on a Random Positioning Machine (RPM) is a new strategy for cartilage regeneration. Using a three-dimensional RPM, a device designed to simulate microgravity on Earth, we investigated the early effects of RPM exposure on human chondrocytes of six different donors after 30 min, 2 h, 4 h, 16 h, and 24 h and compared the results with the corresponding static controls cultured under normal gravity conditions. As little as 30 min of RPM exposure resulted in increased expression of several genes responsible for cell motility, structure and integrity (beta-actin); control of cell growth, cell proliferation, cell differentiation and apoptosis (TGF-ß1, osteopontin); and cytoskeletal components such as microtubules (beta-tubulin) and intermediate filaments (vimentin). After 4 hours of RPM exposure disruptions in the vimentin network were detected. These changes were less dramatic after 16 hours on the RPM, when human chondrocytes appeared to reorganize their cytoskeleton. However, the gene expression and protein content of TGF-ß1 was enhanced during RPM culture for 24 h. Taking these results together, we suggest that chondrocytes exposed to the RPM seem to change their extracellular matrix production behaviour while they rearrange their cytoskeletal proteins prior to forming three-dimensional aggregates.

Ground-based facilities for simulation of microgravity: organism-specific recommendations for their use, and recommended terminology.

Research in microgravity is indispensable to disclose the impact of gravity on biological processes and organisms. However, research in the near-Earth orbit is severely constrained by the limited number of flight opportunities. Ground-based simulators of microgravity are valuable tools for preparing spaceflight experiments, but they also facilitate stand-alone studies and thus provide additional and cost-efficient platforms for gravitational research. The various microgravity simulators that are frequently used by gravitational biologists are based on different physical principles. This comparative study gives an overview of the most frequently used microgravity simulators and demonstrates their individual capacities and limitations. The range of applicability of the various ground-based microgravity simulators for biological specimens was carefully evaluated by using organisms that have been studied extensively under the conditions of real microgravity in space. In addition, current heterogeneous terminology is discussed critically, and recommendations are given for appropriate selection of adequate simulators and consistent use of nomenclature.