Influence of simulated hypogravity on oxygen uptake during treadmill running.

Prolonged exposure to microgravity during spaceflights leads to severe deterioration in the physical performance of astronauts. To understand the effectiveness of existing in-flight daily countermeasures and to plan exercise onboard the International Space Station, we compared supine treadmill running to traditional upright treadmill running on earth. Specifically, we assessed the cardiorespiratory responses to conventional upright running to the responses to supine treadmill running under 0.3 g, 0.6 g, and 1 g of body weight in younger (20-30 years, n = 14, 8 females) and older healthy adults (50-60 years, n = 12, 6 females). Maximal cardiorespiratory capacity was additionally evaluated by performing an incremental running protocol on each treadmill. Maximum speed was greater for 0.3 g and 0.6 g in supine than for upright running (18.5 km/h (1.1) and 15.9 (3.1) vs 13.2 (2.4) p < 0.001). In contrast, maximum oxygen uptake ( V˙O2max ) and maximum heart rate (HRmax ) were greater in upright running than in all supine conditions (Upright treadmill running vs S1.0G vs S0.6G vs S0.3G, 41.7 ml kg-1  min-1 (7.2) vs 30.5 (6.6) vs 32.9 (7.0) vs 30.9 (5.2), p < 0.001 and 171 beats min-1 (14) vs 152 (24) vs 155 (20) vs 152 (18), p < 0.001, respectively). The reduction in V˙O2max was remarkably similar across all three supine conditions, could not be increased by higher running speeds and can be well explained by reduced ground reaction forces (GRF). Thus, although a gravity-related restriction of pulmonary gas exchange or perfusion of the legs when exercising in the supine position can be suspected, findings are also explicable on grounds of the vertical treadmill mechanics. Reduced loading will constitute a substantial limitation to V˙O2 in space with implications for crew health and the physical deterioration of astronauts.

Dragon's Blood Regulates Rac1-WAVE2-Arp2/3 Signaling Pathway to Protect Rat Intestinal Epithelial Barrier Dysfunction Induced by Simulated Microgravity.

Dragon's Blood is a red resin from Dracaena cochinchinensis (Lour.) S.C. Chen (Yunnan, China). As a traditional Chinese medicinal herb, it has shown protective effects on intestinal disorders. Microgravity could alter intestinal homeostasis. However, the potential herbal drugs for preventing intestine epithelial barrier (IEB) dysfunction under microgravity are not available. This study aimed to investigate the effects of Dragon's Blood (DB) on microgravity-induced IEB injury and explore its underlying mechanism. A rat tail-suspension model was used to simulate microgravity (SMG). Histomorphology, ultrastructure, permeability, and expression of junction proteins in jejunum, ileum, and colon of SMG rats were determined. Proteomic analysis was used to identify differentially expressed proteins (DEPs) in rat ileum mucosa altered by DB. The potential mechanism of DB to protect IEB dysfunction was validated by western blotting. The effects of several components in DB were evaluated in SMG-treated Caco-2 cells. DB protected against IEB disruption by repairing microvilli and crypts, inhibiting inflammatory factors, lowering the permeability and upregulating the expression of tight and adherens junction proteins in the ileum of SMG rats. Proteomic analysis showed that DB regulated 1080 DEPs in rat ileum mucosa. DEPs were significantly annotated in cell-cell adhesion, focal adhesion, and cytoskeleton regulation. DB increased the expression of Rac1-WAVE2-Arp2/3 pathway proteins and F-actin to G-actin ratio, which promoted the formation of focal adhesions. Loureirin C in DB showed a protective effect on epithelial barrier injury in SMG-treated Caco-2 cells. DB could protect against IEB dysfunction induced by SMG, and its mechanism is associated with the formation of focal adhesions mediated by the Rac1-WAVE2-Arp2/3 pathway, which benefits intestinal epithelial cell migration and barrier repair.

Simulated Microgravity Suppresses Osteogenic Differentiation of Mesenchymal Stem Cells by Inhibiting Oxidative Phosphorylation.

Studies showed that energy metabolism plays a pivotal role in the differentiation of stem cells. Previous studies revealed that simulated microgravity (SMG) inhibits osteogenic differentiation of mesenchymal stem cells (MSCs). However, the underlying relationship between osteogenesis and energy metabolism under SMG conditions is not fully understood. In the present study, we investigated mitochondrial oxidative phosphorylation (OXPHOS) by assessing the level of peroxisome proliferator activated receptor γ coactivator 1α (PGC-1α), mitochondrial DNA (mtDNA) copy number, mitochondrial mass and oxygen consumption rate (OCR) during osteogenesis of MSCs under SMG conditions. We found that SMG inhibited osteogenic differentiation and OXPHOS of MSCs. Moreover, the expression of sirtuin 1 (Sirt1), an important energy sensor, significantly decreased. After upregulating the expression of Sirt1 using resveratrol, an activator of Sirt1, SMG-inhibited OXPHOS and osteogenic differentiation of MSCs were recovered. Taken together, our results suggest that SMG suppresses osteogenic differentiation of MSCs by inhibiting OXPHOS, indicating that OXPHOS might serve as a potential therapeutic target for repairing bone loss under microgravity conditions.

Simulated microgravity suppresses MAPK pathway-mediated innate immune response to bacterial infection and induces gut microbiota dysbiosis.

During spaceflight, astronauts are subjected to various physical stressors including microgravity, which could cause immune dysfunction and thus potentially predispose astronauts to infections and illness. However, the mechanisms by which microgravity affects innate immunity remain largely unclear. In this study, we conducted RNA-sequencing analysis to show that simulated microgravity (SMG) suppresses the production of inflammatory cytokines including tumor necrosis factor (TNF) and interleukin-6 (IL-6) as well as the activation of the innate immune signaling pathways including the p38 mitogen-activated protein kinase (MAPK) and the Erk1/2 MAPK pathways in the Enteropathogenic escherichia coli (EPEC)-infected macrophage cells. We then adopted hindlimb-unloading (HU) mice, a model mimicking the microgravity of a spaceflight environment, to demonstrate that microgravity suppresses proinflammatory cytokine-mediated intestinal immunity to Citrobacter rodentium infection and induces the disturbance of gut microbiota, both of which phenotypes could be largely corrected by the introduction of VSL#3, a high-concentration probiotic preparation of eight live freeze-dried bacterial species. Taken together, our study provides new insights into microgravity-mediated innate immune suppression and intestinal microbiota disturbance, and suggests that probiotic VSL#3 has great potential as a dietary supplement in protecting individuals from spaceflight mission-associated infections and gut microbiota dysbiosis.

Simulated microgravity disturbs iron metabolism and distribution in humans: Lessons from dry immersion, an innovative ground-based human model.

The objective of the present study was to determine the effects of dry immersion, an innovative ground-based human model of simulated microgravity and extreme physical inactivity, on iron homeostasis and distribution. Twenty young healthy men were recruited and submitted to 5 days of dry immersion (DI). Fasting blood samples and MRI were performed before and after DI exposure to assess iron status, as well as hematological responses. DI increased spleen iron concentrations (SIC), whereas hepatic iron store (HIC) was not affected. Spleen iron sequestration could be due to the concomitant increase in serum hepcidin levels (P < .001). Increased serum unconjugated bilirubin, as well as the rise of serum myoglobin levels support that DI may promote hemolysis and myolysis. These phenomena could contribute to the concomitant increase of serum iron and transferrin saturation levels (P < .001). As HIC remained unchanged, increased serum hepcidin levels could be due both to higher transferrin saturation level, and to low-grade pro-inflammatory as suggested by the significant rise of serum ferritin and haptoglobin levels after DI (P = .003 and P = .003, respectively). These observations highlight the need for better assessment of iron metabolism in bedridden patients, and an optimization of the diet currently proposed to astronauts.

Characterization of C2C12 cells in simulated microgravity: Possible use for myoblast regeneration.

Muscle loss is a major problem for many in lifetime. Muscle and bone degeneration has also been observed in individuals exposed to microgravity and in unloading conditions. C2C12 myoblst cells are able to form myotubes, and myofibers and these cells have been employed for muscle regeneration purposes and in myogenic regeneration and transplantation studies. We exposed C2C12 cells in an random position machine to simulate microgravity and study the energy and the biochemical challenges associated with this treatment. Simulated microgravity exposed C2C12 cells maintain positive proliferation indices and delay the differentiation process for several days. On the other hand this treatment significantly alters many of the biochemical and the metabolic characteristics of the cell cultures including calcium homeostasis. Recent data have shown that these perturbations are due to the inhibition of the ryanodine receptors on the membranes of intracellular calcium stores. We were able to reverse this perturbations treating cells with thapsigargin which prevents the segregation of intracellular calcium ions in the mitochondria and in the sarco/endoplasmic reticula. Calcium homeostasis appear a key target of microgravity exposure. In conclusion, in this study we reported some of the effects induced by the exposure of C2C12 cell cultures to simulated microgravity. The promising information obtained is of fundamental importance in the hope to employ this protocol in the field of regenerative medicine.

Simulated microgravity inhibits C2C12 myogenesis via phospholipase D2-induced Akt/FOXO1 regulation.

The skeletal muscle system has evolved to maintain body posture against a constant gravitational load. Mammalian target of rapamycin (mTOR) regulates the mechanically induced increase in the skeletal muscle mass. In the present study, we investigated mTOR pathway in C2C12 myoblasts in a model of mechanical unloading by creating a simulated microgravity (SM) using 3 D clinorotation. SM decreased the phosphorylation of Akt at Ser 473, which was mediated by mTOR complex 2 (mTORC2), in C2C12 myoblasts, leading to a decrease in the cell growth rate. Subsequently, SM inhibited C2C12 myogenesis in an Akt-dependent manner. In addition, SM increased the phospholipase D (PLD) activity by enhancing PLD2 expression, resulting in the dissociation of mSIN1 from the mTORC2, followed by decrease in the phosphorylation of Akt at Ser 473, and FOXO1 at Ser 256 in C2C12 myoblasts. Exposure to SM decreased the autophagic flux of C2C12 myoblasts by regulation of mRNA level of autophagic genes in a PLD2 and FOXO1-dependent manner, subsequently, resulting in a decrease in the C2C12 myogenesis. In conclusion, by analyzing the molecular signature of C2C12 myogenesis using SM, we suggest that the regulatory axis of the PLD2 induced Akt/FOXO1, is critical for C2C12 myogenesis.

Treatment with hydrogen sulfide donor attenuates bone loss induced by modeled microgravity.

The present study was undertaken to explore the therapeutic potential of hydrogen sulfide against bone loss induced by modeled microgravity. Hindlimb suspension (HLS) and rotary wall vessel bioreactor were applied to model microgravity in vivo and in vitro, respectively. Treatment of rats with GYY4137 (a water soluble donor of hydrogen sulfide, 25 mg/kg per day, i.p.) attenuated HLS-induced reduction of bone mineral density in tibiae, and preserved bone structure in tibiae and mechanical strength in femurs. In HLS group, GYY4137 treatment significantly increased levels of osteocalcin in sera. Interestingly, treatment of HLS rats with GYY4137 enhanced osteoblast surface, but had no significant effect on osteoclast surface of proximal tibiae. In MC3T3-E1 cells exposed to modeled microgravity, GYY4137 stimulated transcriptional levels of runt-related transcription factor 2 and enhanced osteoblastic differentiation, as evidenced by increased mRNA expression and activity of alkaline phosphatase. HLS in rats led to enhanced levels of interleukin 6 in sera, skeletal muscle, and tibiae, which could be attenuated by GYY4137 treatment. Our study showed that GYY4137 preserved bone structure in rats exposed to HLS and promoted osteoblastic differentiation in MC3T3-E1 cells under modeled microgravity.

Short-term effects of simulated microgravity on morphology and gene expression in human breast cancer cells.

INTRODUCTION: Microgravity has been shown to impose various effects on breast cancer cells. We exposed human breast cancer cells to simulated microgravity and studied morphology and alterations in gene expression. MATERIALS AND METHODS: Human breast cancer cells were exposed to simulated microgravity in a random positioning machine (RPM) for 24 h. Morphology was observed under light microscopy, and gene alteration was studied by qPCR. RESULTS: After 24 h, formation of three-dimensional structures (spheroids) occurred. BRCA1 expression was significantly increased (1.9×, p < 0.05) in the adherent cells under simulated microgravity compared to the control. Expression of KRAS was significantly decreased (0.6×, p < 0.05) in the adherent cells compared to the control. VCAM1 was significantly upregulated (6.6×, 2.0×, p < 0.05 each) in the adherent cells under simulated microgravity and in the spheroids. VIM expression was significantly downregulated (0.45×, 0.44×, p < 0.05 each) in the adherent cells under simulated microgravity and in the spheroids. There was no significant alteration in the expression of MAPK1, MMP13, PTEN, and TP53. CONCLUSIONS: Simulated microgravity induces spheroid formation in human breast cancer cells within 24 h and alters gene expression toward modified adhesion properties, enhanced cell repair, and phenotype preservation. Further insights into the underlying mechanisms could open up the way toward new therapies.

Effects of miR-503-5p on apoptosis of human pulmonary microvascular endothelial cells in simulated microgravity.

Recent studies have shown that microRNA (miRNAs) can play important roles in the regulation of endothelial cell (EC) function. However, the expression profile of miRNAs and their effects on the apoptosis of ECs under microgravity conditions remains unclear. In this study, the apoptosis of human pulmonary microvascular endothelial cells (HPMECs) under simulated microgravity was identified by Annexin V and propidium iodide double staining and transmission electron microscopy. miRNA microarray assay was used to screen the differentially expressed miRNAs in HPMECs under simulated microgravity, and eight differentially expressed miRNAs were identified. Specifically, miR-503-5p, which was found to be most significantly upregulated in both microarray and quantitative reverse-transcription polymerase chain reaction assays, was selected for further functional investigation. Overexpression of miR-503-5p induced apoptosis of HPMECs under normal gravity and aggravated the negative effects of simulated microgravity on HPMECs. Furthermore, silencing of miR-503-5p expression effectively attenuated the negative effects of simulated microgravity on HPMECs. Further experiments showed that the mRNA and protein expression of anti-apoptotic factor B-cell lymphoma-2 (Bcl-2), which has been confirmed as a direct target of miR-503-5p, was inhibited by the upregulation of miR-503-5p and increased by the downregulation of miR-503-5p. Taken together, our findings demonstrate, for the first time, that miR-503-5p can induce apoptosis of HPMECs under simulated microgravity through, at least in part, inhibiting the expression of Bcl-2.

Combined Effects of Low-Dose Proton Radiation and Simulated Microgravity on the Mouse Retina and the Hematopoietic System.

The purpose of the current study was to characterize the effects of simulated microgravity and radiation-induced changes in retina and retinal vasculature, and to assess the accompanying early changes in immune cells and hematological parameters. To better understand the effects of spaceflight, we used a combination of treatments designed to simulate both the radiation and low-gravity aspects of space conditions. To simulate the broad energy spectrum of a large solar particle event (SPE) and galactic cosmic ray (GCR) radiation, male C57BL/6J mice were exposed to whole-body irradiation using fully modulated beams of 150-MeV protons containing particles of energy from 0 to 150 MeV and a uniform dose-vs.-depth profile. The mice were also hindlimb-unloaded (HLU) by tail suspension. Mice were unloaded for 7 days, exposed to 50 cGy, unloaded for an additional 7 days and then sacrificed for tissue isolation at days 4 and 30 after the combined treatments. Increases in the number of apoptotic cells were observed in the endothelial cells of mice that received radiation alone or with HLU compared to controls at both days 4 and 30 (P < 0.05). Endothelial nitric oxide synthase (eNOS) levels were significantly elevated in the retina after irradiation only or combined with HLU compared to controls at the 30-day time point (P < 0.05). The most robust changes were observed in the combination group, suggesting a synergistic response to radiation and unloading. For hematopoietic parameters, our analysis indicated the main effects for time and radiation at day 4 after treatments (day 11 postirradiation) (P < 0.05), but a smaller influence of HLU for both white blood cell and lymphocyte counts. The group treated with both radiation and HLU showed greater than 50% reduction in lymphocyte counts compared to controls. Radiation-dependent differences were also noted in specific lymphocyte subpopulations (T, B, natural killer cells). This study shows indications of an early effect of low-dose radiation and spaceflight conditions on retina and immune populations.

Increased Chromosome Aberrations in Cells Exposed Simultaneously to Simulated Microgravity and Radiation.

Space radiation and microgravity (µG) are two major environmental stressors for humans in space travel. One of the fundamental questions in space biology research is whether the combined effects of µG and exposure to cosmic radiation are interactive. While studies addressing this question have been carried out for half a century in space or using simulated µG on the ground, the reported results are ambiguous. For the assessment and management of human health risks in future Moon and Mars missions, it is necessary to obtain more basic data on the molecular and cellular responses to the combined effects of radiation and µG. Recently we incorporated a µG⁻irradiation system consisting of a 3D clinostat synchronized to a carbon-ion or X-ray irradiation system. Our new experimental setup allows us to avoid stopping clinostat rotation during irradiation, which was required in all other previous experiments. Using this system, human fibroblasts were exposed to X-rays or carbon ions under the simulated µG condition, and chromosomes were collected with the premature chromosome condensation method in the first mitosis. Chromosome aberrations (CA) were quantified by the 3-color fluorescent in situ hybridization (FISH) method. Cells exposed to irradiation under the simulated µG condition showed a higher frequency of both simple and complex types of CA compared to cells irradiated under the static condition by either X-rays or carbon ions.

Radio Electric Asymmetric Conveyer (REAC) technology to obviate loss of T cell responsiveness under simulated microgravity.

Alterations of the gravitational environment are likely to modify cell behavior. Several studies have proven that T cells are sensitive to gravity alterations and that microgravity conditions may induce immunosuppression and weakened T cell immune response in humans during spaceflights. The aim of this work was to elucidate if a specific treatment of Radio Electric Asymmetric Conveyer (REAC) technology could restore, after mitogenic activation (Con A), a correct expression of cytokine IL2 gene and its receptor IL2R alpha, which are inhibited in T cells under microgravity conditions, as demonstrated in several studies. The results of this study, conducted in microgravity simulated with Random Positioning Machine (RPM), confirm the T cell activation recovery and offer the evidence that REAC technology could contribute to the understanding of T cell growth responsiveness in space, reducing the impact of weightlessness on the immune system experienced by humans in long duration space missions.

Bone Loss Induced by Simulated Microgravity, Ionizing Radiation and/or Ultradian Rhythms in the Hindlimbs of Rats.

OBJECTIVE: To better understand the pathological causes of bone loss in a space environment, including microgravity, ionizing radiation, and ultradian rhythms. METHODS: Sprague Dawley (SD) rats were randomly divided into a baseline group, a control group, a hindlimb suspension group, a radiation group, a ultradian rhythms group and a combined-three-factor group. After four weeks of hindlimb suspension followed by X-ray exposure and/or ultradian rhythms, biomechanical properties, bone mineral density, histological analysis, microstructure parameters, and bone turnover markers were detected to evaluate bone loss in hindlimbs of rats. RESULTS: Simulated microgravity or combined-three factors treatment led to a significant decrease in the biomechanical properties of bones, reduction in bone mineral density, and deterioration of trabecular parameters. Ionizing radiation exposure also showed adverse impact while ultradian rhythms had no significant effect on these outcomes. Decrease in the concentration of the turnover markers bone alkaline phosphatase (bALP), osteocalcin (OCN), and tartrate-resistant acid phosphatase-5b (TRAP-5b) in serum was in line with the changes in trabecular parameters. CONCLUSION: Simulated microgravity is the main contributor of bone loss. Radiation also results in deleterious effects but ultradian rhythms has no significant effect. Combined-three factors treatment do not exacerbate bone loss when compared to simulated microgravity treatment alone.

Effects of simulated microgravity on human brain nervous tissue.

During spaceflight, the negative effects of space microgravity on astronauts are becoming more and more prominent, and especially, of which on the nervous system is urgently to be solved. For this purpose tissue blocks and primary cells of nervous tissues obtained from glioma of patients were cultivated after culturing for about 7days, explanted tissues and cells were then randomly divided into two groups, one for static culture (control group, C), and the other for rotary processing for 1day, 3days, 5days, 7days and 14days (experiment group, E). Figures captured by inverted microscope revealed that, with short time rotating for 1day or 3days, morphology changes of tissue blocks were not obvious. When the rotary time was extended to 7days or 14days, it was found that cell somas is significantly larger and the ability of adhesion is declined in comparison with that in control group. Additionally, the arrangement of cells migrated from explanted tissues was disorganized, and the migration distance became shorter. In immunofluorescence analysis, ß-tubulin filaments in control group appeared to organize into bundles. While in experiment group, ß-tubulin was highly disorganized. In conclusion, simulated microgravity treatment for a week affected the morphology of nervous tissue, and caused highly disorganized distribution of cytoskeleton and the increase of cell apoptosis. These morphological changes might be one of the causes of apoptosis induced by simulated microgravity.

Melatonin Suppresses Autophagy Induced by Clinostat in Preosteoblast MC3T3-E1 Cells.

Microgravity exposure can cause cardiovascular and immune disorders, muscle atrophy, osteoporosis, and loss of blood and plasma volume. A clinostat device is an effective ground-based tool for simulating microgravity. This study investigated how melatonin suppresses autophagy caused by simulated microgravity in preosteoblast MC3T3-E1 cells. In preosteoblast MC3T3-E1 cells, clinostat rotation induced a significant time-dependent increase in the levels of the autophagosomal marker microtubule-associated protein light chain (LC3), suggesting that autophagy is induced by clinostat rotation in these cells. Melatonin treatment (100, 200 nM) significantly attenuated the clinostat-induced increases in LC3 II protein, and immunofluorescence staining revealed decreased levels of both LC3 and lysosomal-associated membrane protein 2 (Lamp2), indicating a decrease in autophagosomes. The levels of phosphorylation of mammalian target of rapamycin (p-mTOR) (Ser2448), phosphorylation of extracellular signal-regulated kinase (p-ERK), and phosphorylation of serine-threonine protein kinase (p-Akt) (Ser473) were significantly reduced by clinostat rotation. However, their expression levels were significantly recovered by melatonin treatment. Also, expression of the Bcl-2, truncated Bid, Cu/Zn- superoxide dismutase (SOD), and Mn-SOD proteins were significantly increased by melatonin treatment, whereas levels of Bax and catalase were decreased. The endoplasmic reticulum (ER) stress marker GRP78/BiP, IRE1α, and p-PERK proteins were significantly reduced by melatonin treatment. Treatment with the competitive melatonin receptor antagonist luzindole blocked melatonin-induced decreases in LC3 II levels. These results demonstrate that melatonin suppresses clinostat-induced autophagy through increasing the phosphorylation of the ERK/Akt/mTOR proteins. Consequently, melatonin appears to be a potential therapeutic agent for regulating microgravity-related bone loss or osteoporosis.

Scaffold-free Tissue Formation Under Real and Simulated Microgravity Conditions.

Scaffold-free tissue formation in microgravity is a new method in regenerative medicine and an important topic in Space Medicine. In this MiniReview, we focus on recent findings in the field of tissue engineering that were observed by exposing cells to real microgravity in space or to devices simulating to at least some extent microgravity conditions on Earth (ground-based facilities). Under both conditions - real and simulated microgravity - a part of the cultured cells of various populations detaches from the bottom of a culture flask. The cells form three-dimensional (3D) aggregates resembling the organs from which the cells have been derived. As spaceflights are rare and extremely expensive, cell culture under simulated microgravity allows more comprehensive and frequent studies on the scaffold-free 3D tissue formation in some aspects, as a number of publications have proven during the last two decades. In this MiniReview, we summarize data from our own studies and work from various researchers about tissue engineering of multi-cellular spheroids formed by cancer cells, tube formation by endothelial cells and cartilage formation by exposing the cells to ground-based facilities such as the 3D Random Positioning Machine (RPM), the 2D Fast-Rotating Clinostat (FRC) or the Rotating Wall Vessel (RWV). Subsequently, we investigated self-organization of 3D aggregates without scaffolds pursuing to enhance the frequency of 3D formation and to enlarge the size of the organ-like aggregates. The density of the monolayer exposed to real or simulated microgravity as well as the composition of the culture media revealed an impact on the results. Genomic and proteomic alterations were induced by simulated microgravity. Under microgravity conditions, adherent cells expressed other genes than cells grown in spheroids. In this MiniReview, the recent improvements in scaffold-free tissue formation are summarized and relationships between phenotypic and molecular appearance are highlighted.

The combined effects of X-ray radiation and hindlimb suspension on bone loss.

Outer space is a complex environment with various phenomena that negatively affect bone metabolism, including microgravity and highly energized ionizing radiation. In the present study, we used four groups of male Wistar rats treated with or without four-week hindlimb suspension after 4 Gy of X-rays to test whether there is a combined effect for hindlimb suspension and X-ray radiation. We tested trabecular parameters and some cytokines of the bone as leading indicators of bone metabolism. The results showed that hindlimb suspension and X-ray radiation could cause a significant increase in bone loss. Hindlimb suspension caused a 56.6% bone loss (P = 0.036), while X-ray radiation caused a 30.7% (P = 0.041) bone loss when compared with the control group. The combined factors of hindlimb suspension and X-rays exerted a combined effect on bone mass, with a reduction of 64.8% (P = 0.003).

21 Days head-down bed rest induces weakening of cell-mediated immunity - Some spaceflight findings confirmed in a ground-based analog.

Several studies indicate a weakening of cell-mediated immunity (CMI) and reactivation of latent herpes viruses during spaceflight. We tested the hypothesis that head-down bed rest (HDBR), a ground-based analog of spaceflight, mimics the impact of microgravity on human immunity. Seven healthy young males underwent two periods of 3 weeks HDBR in the test facility of the German Aerospace Center. As a nutritional countermeasure aimed against bone demineralisation, 90 mmol potassium bicarbonate (KHCO(3)) was administered daily in a crossover design. Blood samples were drawn on five occasions. Whole blood was stimulated with antigen i.e. Candida albicans, purified protein derivative (PPD) tuberculin, tetanus toxoid and Cytomegalovirus (CMV) (CMV-QuantiFERON). Flow cytometric analysis included CD4(+)CD25(+)CD127(-)FOXP3(+) regulatory T cells (Tregs), γδ T cells, B cells, NK cells and dendritic cells. In one of the two bed rest periods, we observed a significant decrease in production of interleukin-2 (IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) following phytohemagglutinin (PHA) stimulation, with a rapid normalization being observed after HDBR. The cytokine levels showed a V-shaped pattern that led to a relativeTh2-shift in cytokine balance. Only three individuals responded to the specific T cell antigens without showing signs of an altered response during HDBR, nor did we observe reactivation of CMV or Epstein-Barr virus (EBV). Of unknown significance, dietary supplementation with KHCO(3) counteracted the decrease in IL-2 levels during HDBR, while there was no impact on other immunological parameters. We conclude that discrete alterations in CMI may be induced by HDBR in selected individuals.

The effect of microgravity on tissue structure and function of rat testis

To explore whether an environment of weightlessness will cause damage to the reproductive system of animals, we used the tail-suspension model to simulate microgravity, and investigated the effect of microgravity on the tissue structure and function of the testis in sexually mature male rats. Forty-eight male Wistar rats weighing 200-250 g were randomly assigned to three groups (N = 16 each): control, tail traction, and tail suspension. After the rats were suspended for 7 or 14 days, morphological changes of testis were evaluated by histological and electron microscopic methods. The expression of HSP70, bax/bcl-2 and AR (androgen receptor) in testis was measured by immunohistochemistry. Obvious pathological lesions were present in the testis after the rats were suspended for 7 or 14 days. We detected overexpression of HSP70 and an increase of apoptotic cells, which may have contributed to the injury to the testis. The expression of AR, as an effector molecule in the testis, was significantly decreased in the suspended groups compared to control (P < 0.01). We also observed that, with a longer time of suspension, the aforementioned pathological damage became more serious and some pathological injury to the testis was irreversible. The results demonstrated that a short- or medium-term microgravity environment could lead to severe irreversible damage to the structure of rat testis.