PCR-based analysis of microbial communities during the EuroGeoMars campaign at Mars Desert Research Station, Utah

The search for evidence of past or present life on Mars will require the detection of markers that indicate the presence of life. Because deoxyribonucleic acid (DNA) is found in all known living organisms, it is considered to be a ‘biosignature' of life. The main function of DNA is the long-term storage of genetic information, which is passed on from generation to generation as hereditary material. The Polymerase Chain Reaction (PCR) is a revolutionary technique which allows a single fragment or a small number of fragments of a DNA molecule to be amplified millions of times, making it possible to detect minimal traces of DNA. The compactness of the contemporary PCR instruments makes routine sample analysis possible with a minimum amount of laboratory space. Furthermore the technique is effective, robust and straightforward. Our goal was to establish a routine for the detection of DNA from micro-organisms using the PCR technique during the EuroGeoMars simulation campaign. This took place at the Mars Society's Mars Desert Research Station (MDRS) in Utah in February 2009 (organized with the support of the International Lunar Exploration Working Group (ILEWG), NASA Ames and the European Space Research and Technology Centre (ESTEC)). During the MDRS simulation, we showed that it is possible to establish a minimal molecular biology lab in the habitat for the immediate on-site analysis of samples by PCR after sample collection. Soil and water samples were taken at different locations and soil depths. The sample analysis was started immediately after the crew returned to the habitat laboratory. DNA was isolated from micro-organisms and used as a template for PCR analysis of the highly conserved ribosomal DNA to identify representatives of the different groups of micro-organisms (bacteria, archaea and eukarya). The PCR products were visualized by agarose gel electrophoresis and documented by transillumination and digital imaging. The microbial diversity in the collected samples was analysed with respect to sampling depth and the presence or absence of vegetation. For the first time, we have demonstrated that it is possible to perform direct on-site DNA analysis by PCR at MDRS, a simulated planetary habitat in an extreme environment that serves as a model for preparation and optimization of techniques to be used for future Mars exploratio

Cytoskeletal stability and metabolic alterations in primary human macrophages in long-term microgravity

The immune system is one of the most affected systems of the human body during space flight. The cells of the immune system are exceptionally sensitive to microgravity. Thus, serious concerns arise, whether space flight associated weakening of the immune system ultimately precludes the expansion of human presence beyond the Earth's orbit. For human space flight, it is an urgent need to understand the cellular and molecular mechanisms by which altered gravity influences and changes the functions of immune cells. The CELLBOX-PRIME (= CellBox-Primary Human Macrophages in Microgravity Environment) experiment investigated for the first time microgravity-associated long-term alterations in primary human macrophages, one of the most important effector cells of the immune system. The experiment was conducted in the U.S. National Laboratory on board of the International Space Station ISS using the NanoRacks laboratory and Biorack type I standard CELLBOX EUE type IV containers. Upload and download were performed with the SpaceX CRS-3 and the Dragon spaceship on April 18th, 2014 / May 18th, 2014. Surprisingly, primary human macrophages exhibited neither quantitative nor structural changes of the actin and vimentin cytoskeleton after 11 days in microgravity when compared to 1g controls. Neither CD18 or CD14 surface expression were altered in microgravity, however ICAM-1 expression was reduced. The analysis of 74 metabolites in the cell culture supernatant by GC-TOF-MS, revealed eight metabolites with significantly different quantities when compared to 1g controls. In particular, the significant increase of free fucose in the cell culture supernatant was associated with a significant decrease of cell surface-bound fucose. The reduced ICAM-1 expression and the loss of cell surface-bound fucose may contribute to functional impairments, e.g. the activation of T cells, migration and activation of the innate immune response. We assume that the surprisingly small and non-significant cytoskeletal alterations represent a stable "steady state" after adaptive processes are initiated in the new microgravity environment. Due to the utmost importance of the human macrophage system for the elimination of pathogens and the clearance of apoptotic cells, its apparent robustness to a low gravity environment is crucial for human health and performance during long-term space missions

Cytoskeletal stability and metabolic alterations in primary human macrophages in long-term microgravity

The immune system is one of the most affected systems of the human body during space flight. The cells of the immune system are exceptionally sensitive to microgravity. Thus, serious concerns arise, whether space flight associated weakening of the immune system ultimately precludes the expansion of human presence beyond the Earth's orbit. For human space flight, it is an urgent need to understand the cellular and molecular mechanisms by which altered gravity influences and changes the functions of immune cells. The CELLBOXPRIME (= CellBox-Primary Human Macrophages in Microgravity Environment) experiment investigated for the first time microgravity-associated long-term alterations in primary human macrophages, one of the most important effector cells of the immune system. The experiment was conducted in the U. S. National Laboratory on board of the International Space Station ISS using the NanoRacks laboratory and Biorack type I standard CELLBOX EUE type IV containers. Upload and download were performed with the SpaceX CRS- 3 and the Dragon spaceship on April 18th, 2014 / May 18th, 2014. Surprisingly, primary human macrophages exhibited neither quantitative nor structural changes of the actin and vimentin cytoskeleton after 11 days in microgravity when compared to 1g controls. Neither CD18 or CD14 surface expression were altered in microgravity, however ICAM-1 expression was reduced. The analysis of 74 metabolites in the cell culture supernatant by GC-TOF-MS, revealed eight metabolites with significantly different quantities when compared to 1g controls. In particular, the significant increase of free fucose in the cell culture supernatant was associated with a significant decrease of cell surface-bound fucose. The reduced ICAM-1 expression and the loss of cell surface-bound fucose may contribute to functional impairments, e.g. the activation of T cells, migration and activation of the innate immune response. We assume that the surprisingly small and non-significant cytoskeletal alterations represent a '' stable-steady state '' after adaptive processes are initiated in the new microgravity environment. Due to the utmost importance of the human macrophage system for the elimination of pathogens and the clearance of apoptotic cells, its apparent robustness to a low gravity environment is crucial for human health and performance during long-term space missions

Amyloid Precursor Protein Is Trafficked and Secreted via Synaptic Vesicles

A large body of evidence has implicated amyloid precursor protein (APP) and its proteolytic derivatives as key players in the physiological context of neuronal synaptogenesis and synapse maintenance, as well as in the pathology of Alzheimer's Disease (AD). Although APP processing and release are known to occur in response to neuronal stimulation, the exact mechanism by which APP reaches the neuronal surface is unclear. We now demonstrate that a small but relevant number of synaptic vesicles contain APP, which can be released during neuronal activity, and most likely represent the major exocytic pathway of APP. This novel finding leads us to propose a revised model of presynaptic APP trafficking that reconciles existing knowledge on APP with our present understanding of vesicular release and recycling

Schnelle zelluläre Reaktion und Anpassung an Schwerelosigkeit

Zelluläre Prozesse werden durch Änderungen der Gravitationskraft in vielfältiger Weise beeinflusst. In unseren Studien konnten wir in verschiedenen zellulären Systemen und in verschiedenen Forschungsplattformen (2D Klinostat, Parabelflüge, suborbitale Forschungsraketen, Internationale Raumstation) nachweisen, dass Reaktionen und Anpassungsprozesse bereits innerhalb von Sekunden bis Minuten nach Beginn der veränderten Schwerkraft auftreten. Diese schnellen Prozesse fanden sich in der Genexpression, der Zellzykluskontrolle, der Signaltransduktion, der Zytoskelettorganisation und des oxidativen Burst. Es stellt sich die Frage nach der Ursache der Transduktion einer unspezifischen Kraft in eine hochspezifische zelluläre Antwort, die in der Spezifität der zellulären Eigenschaften begründet liegen kann. Es ist denkbar, dass die Gravitationskraft der Erde die Chromatinarchitektur und deren Zugänglichkeit stabilisiert und eine homöostatische Bedingung für die Genexpression darstellt. Die Untersuchung mechanobiologischer Mechanismen der Genexpression in Schwerelosigkeit kann die Aufdeckung fundamentaler Prinzipien ermöglichen, wie mechanische Kräfte die Zellfunktion regulieren

Hochauflösende Fluoreszenzmikroskopie in lebenden Zellen des Immunsystems unter Raumflugbedingungen

Zelluläre Prozesse werden durch Änderungen der Gravitationskraft in vielfältiger Weise dauerhaft beeinflusst. Die Möglichkeit der kontinuierlichen Beobachtung dieser Abläufe ist von großer Relevanz. Die Etablierung und Anwendung von hochauflösender Fluoreszenzmikroskopie im Rahmen des FLUMIAS-Experiments erlaubte kontinuierliche Aufnahmen von lebenden Immunzellen an Bord der Forschungsraketenmission TEXUS-54. Gravitationsbedingte Kurzzeitänderungen der Zellmorphologie sowie des Zellskeletts und assoziierte schnelle zelluläre Anpassungsvorgänge konnten in primären menschlichen Makrophagen identifiziert werden. Die hier vorgestellte Möglichkeit der Echtzeitanalyse kann das Wissen über dynamische zelluläre Reaktionen und Adaptationsprozesse an die Weltraumbedingungen enorm erweitern

Time course of cellular and molecular regulation in the immune system in altered gravity: progressive damage or adaptation?

We summarized the current knowledge about adaptation processes of isolated immune cells, animal models and the human body to altered gravity conditions. Many studies indicate an adaptation reaction of the immune system to the new microgravity environment, at least for the T cell system. Animal and human studies indicated adaptation processes starting after two weeks and continuing until 6 month or longer, which was reflected by cytokine concentrations in blood plasma or in stimulation assays. Adaptive reactions regarding IFN-c, TNF-a and IL-2 concentrations were detected after 12 days spaceflight in animal studies and after 2–4 months in human studies, whereas adaptive reactions regarding IL-4, IL-6, IL-8 and IL-10 were found after 6 months spaceflight. Cellular studies were performed mainly as short-term studies, and only a few studies addressed alterations longer than 3 days. However, cross validation between studies is often not possible or indicated conflicting results. Many in vitro studies, mostly done with T lymphocytes, demonstrated extensive cellular and molecular alterations. In contrast, long-term studies with animals and humans are completely lacking this dramatic picture of short-term cellular effects, which indicates a very efficient adaptation process, partially evidenced by new steady state of adaptive response in the human immune system after weeks until months. Therefore, we assume that the human body and its cells are equipped with a robust and efficient adaptation potential when challenged with low gravitational environments