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The immune system belongs to the most affected systems during spaceflight, and sensitivity of cells of the human immune system to reduced gravity has been confirmed in numerous studies in real and simulated microgravity. Immune system dysfunction during spaceflight represents a substantial risk for exploration class mission knowledge about the clinical, cellular, and genetic basis of immune system response, and adaptation to altered gravity will provide key information for appropriate risk management, efficient monitoring, and countermeasures against existing limiting factors for human health and performance during manned exploration of the solar system. In spite of the immune system dysregulation, studies indicate an adaptation reaction of the immune system to the new microgravity environment, at least for the T-cell system, starting after 2 weeks and continuing until 6 months or longer, reflected by cytokine concentrations in blood plasma or in stimulation assays. At the cellular level, rapid adaptation responses could be detected as early as after seconds until minutes in T cells and macrophages. Therefore, adaptive responses of cells and the whole organism could be expected under microgravity and altered gravity in general. Preventive countermeasures should therefore consider support and stabilization of the endogenous adaptation programs. Potential countermeasures for risk mitigation are summarized in this chapter. We assume that the immune systems not only have a significant adaptation potential when challenged with low gravitational environments but also provide interesting preventive and therapeutic options for long-term space missions.

BACKGROUND: Injury to the lateral femoral cutaneous nerve (LFCN) is a risk during the operative anterior approach to the hip joint. Although several anatomical studies have described the proximal course of the nerve in relation to the anterior superior iliac spine (ASIS) and the inguinal ligament, the distal course of the LFCN in the proximal aspect of the thigh has not been sufficiently studied. The aim of this cadaveric study was to examine the branching pattern of the nerve, with special consideration to the anterior approach to the hip joint. METHODS: Twenty-eight cadaveric hemipelves from 18 donors (10 paired and 8 unpaired specimens) were dissected. The LFCN branches were localized proximal to the inguinal ligament and traced distally into the area of the proximal aspect of the thigh. Distribution patterns of the nerve with respect to its relationship to the ASIS and the internervous plane of the anterior approach to the hip joint were recorded. RESULTS: We found 3 different branching patterns of the LFCN: sartorius-type (in 36% of the specimens), characterized by a dominant anterior nerve branch coursing along the lateral border of the sartorius muscle with no, or only a thin, posterior branch; posterior-type (in 32%), characterized by a strong posterior nerve branch; and fan-type (in 32%), characterized by multiple spreading nerve branches of equal thickness. In 50% of the specimens, the LFCN divided into ≥2 branches superior to the inguinal ligament. Sixty-two percent of the LFCN branches entered the proximal aspect of the thigh medial to the ASIS; 27%, above; and 11%, lateral to the ASIS. The LFCN consistently coursed within the deep layer of the subcutaneous fat tissue. CONCLUSIONS: Injury to branches of the LFCN cannot be avoided in approximately one-third of surgical dissections that use the anterior approach to the hip joint. To protect the anterior branch of the LFCN, the skin incision should be as lateral as possible. The posterior branch of the LFCN

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Gravity has been a constant force throughout the Earth's evolutionary history. Thus, one of the fundamental biological questions is if and how complex cellular and molecular functions of life on Earth require gravity. In this study, we investigated the influence of gravity on the oxidative burst reaction in macrophages, one of the key elements in innate immune response and cellular signaling. An important step is the production of superoxide by the NADPH oxidase, which is rapidly converted to H2O2 by spontaneous and enzymatic dismutation. The phagozytosis-mediated oxidative burst under altered gravity conditions was studied in NR8383 rat alveolar macrophages by means of a luminol assay. Ground-based experiments in "functional weightlessness" were performed using a 2 D clinostat combined with a photomultiplier (PMT clinostat). The same technical set-up was used during the 13th DLR and 51st ESA parabolic flight campaign. Furthermore, hypergravity conditions were provided by using the Multi-Sample Incubation Centrifuge (MuSIC) and the Short Arm Human Centrifuge (SAHC). The results demonstrate that release of reactive oxygen species (ROS) during the oxidative burst reaction depends greatly on gravity conditions. ROS release is 1.) reduced in microgravity, 2.) enhanced in hypergravity and 3.) responds rapidly and reversible to altered gravity within seconds. We substantiated the effect of altered gravity on oxidative burst reaction in two independent experimental systems, parabolic flights and 2D clinostat / centrifuge experiments. Furthermore, the results obtained in simulated microgravity (2D clinorotation experiments) were proven by experiments in real microgravity as in both cases a pronounced reduction in ROS was observed. Our experiments indicate that gravity-sensitive steps are located both in the initial activation pathways and in the final oxidative burst reaction itself, which could be explained by the role of cytoskeletal dynamics in the assembly and function

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.

Sensitivity of the human immune system to microgravity has been supposed since the first Apollo missions and was demonstrated during several space missions in the past. In vitro experiments demonstrated that cells of the immune system are exceptionally sensitive to microgravity. Therefore, serious concerns arose whether spaceflight-associated immune system weakening ultimately precludes the expansion of human presence beyond Earth’s orbit. In human cells, gravitational forces may be sensed by an individual cell in the context of altered extracellular matrix mechanics, cell shape, cytoskeletal organization, or internal prestress in the cell–tissue matrix. The development of cellular mechanosensitivity and signal transduction was probably an evolutionary requirement to enable our cells to sense their individual microenvironment. Therefore it is possible that the same mechanisms, which enable human cells to sense and to cope with mechanical stress, are potentially dangerous in microgravity.

In this study we investigated if and how cannabinoid receptor stimulation regulates macrophageal differentiation, which is one of the key steps in the immune effector reaction. For that reason, we used a well established differentiation model system of human U937 myelocytic leukemia cells that differentiate along the monocyte/macrophage lineage upon stimulation with the phorbol ester PMA. Constant cannabinoid receptor (CB) stimulation was performed using WIN55212-2, a potent synthetic CB agonist. We found that WIN55212-2 inhibited CB1/2-receptor-dependent PMA-induced differentiation of human myelocytic U937 cells into the macrophageal phenotype, which was associated with impaired vimentin, ICAM-1 and CD11b expression. In the presence of WIN55212-2, cdc2 protein and mRNA expression was progressively enhanced and Tyr-15-phosporylation of cdc2 was reduced in differentiating U937 cells. Additionally, p21Waf1/Cip1 expression was up-regulated. PMA-induced apoptosis was not enhanced by WIN55212-2 and differentiation-associated c-jun expression was not altered. In conclusion, we suppose that WIN55212-2-induced signals interferes with cell-cycle-arrest-signaling in differentiating myelocytic cells and thus inhibits macrophageal differentiation. Thus, it is possible that the cannabinoid system is able to influence one of the key steps in the immune effector function, the monocytic-macrophageal differentiation by alteration of cell cycle control proteins cdc2 and p21, and is therefore representing a promising option for therapeutic intervention in exacerbated immune reactions.

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 samp

BACKGROUND: Adult neurogenesis is a particular example of brain plasticity that is partially modulated by the endocannabinoid system. Whereas the impact of synthetic cannabinoids on the neuronal progenitor cells has been described, there has been lack of information about the action of plant-derived extracts on neurogenesis. Therefore we here focused on the effects of Delta9-tetrahydrocannabinol (THC) and Cannabidiol (CBD) fed to female C57Bl/6 and Nestin-GFP-reporter mice on proliferation and maturation of neuronal progenitor cells and spatial learning performance. In addition we used cannabinoid receptor 1 (CB1) deficient mice and treatment with CB1 antagonist AM251 in Nestin-GFP-reporter mice to investigate the role of the CB1 receptor in adult neurogenesis in detail. RESULTS: THC and CBD differed in their effects on spatial learning and adult neurogenesis. CBD did not impair learning but increased adult neurogenesis, whereas THC reduced learning without affecting adult neurogenesis. We found the neurogenic effect of CBD to be dependent on the CB1 receptor, which is expressed over the whole dentate gyrus. Similarly, the neurogenic effect of environmental enrichment and voluntary wheel running depends on the presence of the CB1 receptor. We found that in the absence of CB1 receptors, cell proliferation was increased and neuronal differentiation reduced, which could be related to CB1 receptor mediated signaling in Doublecortin (DCX)-expressing intermediate progenitor cells. CONCLUSION: CB1 affected the stages of adult neurogenesis that involve intermediate highly proliferative progenitor cells and the survival and maturation of new neurons. The pro-neurogenic effects of CBD might explain some of the positive therapeutic features of CBD-based compounds.

Microglial cells maintain the immunological integrity of the healthy brain and can exert protection from traumatic injury. During ischemic tissue damage such as stroke, peripheral immune cells acutely infiltrate the brain and may exacerbate neurodegeneration. Whether and how microglia can protect from this insult is unknown. Polymorphonuclear neutrophils (PMNs) are a prominent immunologic infiltrate of ischemic lesions in vivo. Here, we show in organotypic brain slices that externally applied invading PMNs massively enhance ischemic neurotoxicity. This, however, is counteracted by additional application of microglia. Time-lapse imaging shows that microglia exert protection by rapid engulfment of apoptotic, but, strikingly, also viable, motile PMNs in cell culture and within brain slices. PMN engulfment is mediated by integrin- and lectin-based recognition. Interference with this process using RGDS peptides and N-acetyl-glucosamine blocks engulfment of PMNs and completely abrogates the neuroprotective function of microglia. Thus, engulfment of invading PMNs by microglia may represent an entirely new mechanism of CNS immune privilege.

Life on Earth developed in the presence and under the constant influence of gravity. Gravity has been present during the entire evolution, from the first organic molecule to mammals and humans. Modern research revealed clearly that gravity is important, probably indispensable for the function of living systems, from unicellular organisms to men. Thus, gravity research is no more or less a fundamental question about the conditions of life on Earth. Since the first space missions and supported thereafter by a multitude of space and ground-based experiments, it is well known that immune cell function is severely suppressed in microgravity, which renders the cells of the immune system an ideal model organism to investigate the influence of gravity on the cellular and molecular level. Here we review the current knowledge about the question, if and how cellular signal transduction depends on the existence of gravity, with special focus on cells of the immune system. Since immune cell function is fundamental to keep the organism under imnological surveillance during the defence against pathogens, to investigate the effects and possible molecular mechanisms of altered gravity is indispensable for long-term space flights to Earth Moon or Mars. Thus, understanding the impact of gravity on cellular functions on Earth will provide not only important informations about the development of life on Earth, but also for therapeutic and preventive strategies to cope successfully with medical problems during space exploration.

Publisher Summary This chapter describes the permeabilized cell system and the methods used to study the role of Rab-GDI in the reversible membrane association of Rab proteins. The advantage of using permeabilized cells is that, while the cellular organization is maintained, the intracellular membranes are accessible to biochemical manipulations. Thus, this in vitro system closely mimics the physiological conditions in intact cells. Rab proteins are associated with specific compartments along the exocytic and endocytic pathways. Membrane association requires posttranslational modification of the C-terminal cysteines by the addition of geranylgeranyl moieties. In addition, Rab proteins are also present in the cytosol. Cytosolic Rab proteins are complexed to Rab-GDI (GDP dissociation inhibitor), a factor that inhibits the dissociation of GDP and removes Rab proteins from the membrane. These biochemical properties have led to the suggestion that Rab-GDI allows Rab proteins to shuttle between the membrane and the cytosol to serve multiple rounds of vesicular traffic.