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Integrated biochemical and biophysical signals regulate embryonic development. Correct neural tube formation is critical for the development of central nervous system. However, the role of microgravity in neurodevelopment and its underlying molecular mechanisms remain unclear. In this study, the effects of stimulated microgravity (SMG) on the development of human brain organoids are investigated. SMG impairs N-cadherin-based adherens junction formation, leading to neural tube defects associated with dysregulated self-renewal capacity and neuroepithelial disorganization in human brain organoids. Bulk gene expression analyses reveal that SMG alters Hippo and BMP signaling in brain organoids. The neuropathological deficits in SMG-treated organoids can be rescued by regulating YAP/BMP/ID1 axis. Furthermore, sing-cell RNA sequencing data show that SMG results in perturbations in the number and function of neural stem and progenitor cell subpopulations. One of these subpopulations senses SMG cues and transmits BMP signals to the subpopulation responsible for tube morphogenesis, ultimately affecting the proliferating cell population. Finally, SMG intervention leads to persistent neurologic damage even after returning to normal gravity conditions. Collectively, this study reveals molecular and cellular abnormalities associated with SMG during human brain development, providing opportunities for countermeasures to maintain normal neurodevelopment in space., (© 2024 The Author(s). Advanced Science published by Wiley‐VCH GmbH.)

Spaceflight-induced multi-organ dysfunction affects the health of astronauts and the safety of in-orbit flight. However, the effect of microgravity on the kidney and the underlying mechanisms are unknown. In the current study, we used a hindlimb unweighting (HU) animal model to simulate microgravity and employed histological analysis, ischemia-reperfusion experiments, renal ultrasonography, bioinformatics analysis, isometric force measurement, and other molecular experimental settings to evaluate the effects of microgravity on the kidneys and the underlying mechanisms involved in this transition. Relative to controls, 31-day hindlimb unweighting had no obvious effect on serum creatinine and urea nitrogen levels as well as on renal injury scores; however, animals in the HU group showed an impaired renal recovery to ischemia-reperfusion injury. Ultrasonography showed that renal resistive index was increased, which indicated an altered renal hemodynamics induced by simulated microgravity. The enhanced vasoconstriction mediated by the Rho-kinase pathway and impaired vasodilation mediated by NO-eNOS of the intrarenal artery contributed to the altered renal hemodynamics. Simulated microgravity predisposes the kidney to ischemia-reperfusion injury. The altered renal hemodynamics induced by renal arterial remodeling may account for the increased renal injury susceptibility, providing a target for early intervention in kidney dysfunction under microgravity., (© 2025 Federation of American Societies for Experimental Biology.)

Research into the mechanisms by which gravity influences spermatozoa has implications for maintaining the species in deep space exploration and may provide new approaches to reproductive technologies on Earth. Changes in the speed of mouse spermatozoa after 30 min exposure to simulated weightlessness (by 3D-clinostat) and 2 g hypergravity (by centrifugation) were studied using inhibitory analysis. Simulated microgravity after 30 min led to an increase in the speed of spermatozoa and against the background of an increase in the relative calcium content in the cytoplasm. This effect was prevented by the introduction of 6-(dimethylamino) purine, wortmannin, and calyculin A. Hypergravity led to a decrease in the speed of spermatozoa movement, which was prevented by sodium orthovanadate and calyculin A. At the same time, under microgravity conditions, there was a redistribution of proteins forming microfilament bundles between the membrane and cytoplasmic compartments and under hypergravity conditions-proteins forming networks. The obtained results indicate that even a short exposure of spermatozoa to altered gravity leads to the launch of mechanotransduction pathways in them and a change in motility.

The development of osteopenia is one of the most noticeable manifestations of the adverse effects of space factors on crew members. The Hippo signaling pathway has been shown to play a central role in regulating the functional activity of cells through their response to mechanical stimuli. In the present study, the components of the Hippo pathway and the protective properties of osteodifferentiation inducers were investigated under simulated microgravity (smg) using a heterotypic bone marrow cell culture model, which allows for the maintenance of the close interaction between the stromal and hematopoietic compartments, present in vivo and of great importance for both the fate of osteoprogenitors and hematopoiesis. After 14 days of smg, the osteopotential and osteodifferentiation of bone marrow stromal progenitor cells, the expression of Hippo cascade genes and the immunocytochemical status of the adherent fraction of bone marrow cells, as well as the paracrine profile in the conditioned medium and the localization of Yap1 and Runx2 in mechanosensitive cells of the bone marrow were obtained. Simulated microgravity negatively affects stromal and hematopoietic cells when interacting in a heterotypic murine bone marrow cell culture. This is evidenced by the decrease in cell proliferation and osteopotential. Changes in the production of pleiotropic cytokines IL-6, GROβ and MCP-1 were revealed. Fourteen days of simulated microgravity induced a decrease in the nuclear translocation of Yap1 and the transcription factor Runx2 in the stromal cells of the intact group. Exposure to osteogenic induction conditions partially compensated for the negative effect of simulated microgravity. The data obtained will be crucial for understanding the effects of spaceflight on osteoprogenitor cell growth and differentiation via Hippo-Yap signaling.

Background: Orthostatic intolerance occurs following immobilization in patients on Earth and in astronauts after spaceflight. Head-down tilt bedrest is a terrestrial model for weightlessness and induces orthostatic intolerance. We hypothesized that lower-body negative pressure (LBNP) or cycling followed by wearing venous constrictive thigh cuffs mitigates orthostatic intolerance after head-down tilt bedrest., Methods and Results: We enrolled 47 healthy individuals (20 women, 35±9 years) to a 30-day strict head-down tilt bedrest study. During bedrest, they were assigned to 6 hours of 25 mm Hg LBNP (n=12) per day and 1 hour of supine cycling followed by 6 hours of venous constriction through thigh cuffs 6 days per week (n=12), 6 hours of daily upright sitting (positive control, n=11), or no countermeasure (negative control, n=12). We measured orthostatic tolerance as the time to presyncope during 80° head-up tilt testing with incremental LBNP before and immediately after bedrest. We determined plasma volume with carbon monoxide rebreathing before and at the end of bedrest. After bedrest, orthostatic tolerance decreased 540±457 seconds in the control group, 539±68 seconds in the cycling group, 217±379 seconds in the LBNP group, and 289±89 seconds in the seated group ( P <0.0001 time point, P =0.009 for group differences). Supine and upright heart rate increased in all groups following bedrest. Plasma volume was only maintained in the cycling group but decreased in all others (interaction countermeasure×time point P <0.0001)., Conclusions: Six hours of moderate LBNP training was as effective as sitting in attenuating orthostatic intolerance after 30 days of head-down tilt bedrest. Daily cycling exercise followed by 6 hours of wearing venous constrictive thigh cuffs, while maintaining plasma volume, did not improve orthostatic tolerance., Registration: URL: https://www.bfarm.de/EN; Identifiers: DRKS00027643 and DRKS00030848.

Background: Bone loss caused by microgravity exposure presents a serious threat to the health of astronauts, but existing treatment strategies have specific restrictions. This research aimed to investigate whether salidroside (SAL) can mitigate microgravity-induced bone loss and its underlying mechanism., Methods: In this research, we used hindlimb unloading (HLU) and the Rotary Cell Culture System (RCCS) to imitate microgravity in vivo and in vitro., Results: The results showed that salidroside primarily enhances bone density, microstructure, and biomechanical properties by stimulating bone formation and suppressing bone resorption, thereby preserving bone mass in HLU rats. In MC3T3-E1 cells cultured under simulated microgravity in rotary wall vessel bioreactors, the expression of osteogenic genes significantly increased after salidroside administration, indicating that salidroside can promote osteoblast differentiation under microgravity conditions. Furthermore, the Nrf2 inhibitor ML385 diminished the therapeutic impact of salidroside on microgravity-induced bone loss. Overall, this research provides the first evidence that salidroside can mitigate bone loss induced by microgravity exposure through stimulating the Nrf2/HO-1 pathway., Conclusion: These findings indicate that salidroside has great potential for treating space-related bone loss in astronauts and suggest that Nrf2/HO-1 is a viable target for counteracting microgravity-induced bone damage., (© 2024. The Author(s).)

Exposure to the space microenvironment has been found to disrupt the homeostasis of intestinal epithelial cells and alter the composition of the microbiota. To investigate this in more detail and to examine the impact of ginsenoside Rb1, we utilized a mouse model of hindlimb unloading (HU) for four weeks to simulate the effects of microgravity. Our findings revealed that HU mice had ileum epithelial injury with a decrease in the number of intestinal stem cells (ISCs) and the level of cell proliferation. The niche functions for ISCs were also impaired in HU mice, including a reduction in Paneth cells and Wnt signaling, along with an increase in oxidative stress. The administration of Rb1 during the entire duration of HU alleviated the observed intestinal defects, suggesting its beneficial influence on epithelial cell homeostasis. Hindlimb unloading also resulted in gut dysbiosis. The supplementation of Rb1 in the HU mice or the addition of Rb1 derivative compound K in bacterial culture in vitro promoted the growth of beneficial probiotic species such as Akkermansia . The co-housing experiment further showed that Rb1 treatment in ground control mice alone could alleviate the defects in HU mice that were co-housed with Rb1-treated ground mice. Together, these results underscore a close relationship between dysbiosis and impaired ISC functions in the HU mouse model. It also highlights the beneficial effects of Rb1 in mitigating HU-induced epithelial injury by promoting the expansion of intestinal probiotics. These animal-based insights provide valuable knowledge for the development of improved approaches to maintaining ISC homeostasis in astronauts.

Missions into Deep Space are planned this decade. Yet the health consequences of exposure to microgravity and galactic cosmic radiation (GCR) over years-long missions on indispensable visceral organs such as the kidney are largely unexplored. We performed biomolecular (epigenomic, transcriptomic, proteomic, epiproteomic, metabolomic, metagenomic), clinical chemistry (electrolytes, endocrinology, biochemistry) and morphometry (histology, 3D imaging, miRNA-ISH, tissue weights) analyses using samples and datasets available from 11 spaceflight-exposed mouse and 5 human, 1 simulated microgravity rat and 4 simulated GCR-exposed mouse missions. We found that spaceflight induces: 1) renal transporter dephosphorylation which may indicate astronauts' increased risk of nephrolithiasis is in part a primary renal phenomenon rather than solely a secondary consequence of bone loss; 2) remodelling of the nephron that results in expansion of distal convoluted tubule size but loss of overall tubule density; 3) renal damage and dysfunction when exposed to a Mars roundtrip dose-equivalent of simulated GCR., (© 2024. The Author(s).)

Muscle wasting is a major problem leading to reduced quality of life and higher risks of mortality and various diseases. Muscle atrophy is caused by multiple conditions in which protein degradation exceeds its synthesis, including disuse, malnutrition, and microgravity. While Vitamin D receptor (VDR) is well known to regulate calcium and phosphate metabolism to maintain bone, recent studies have shown that VDR also plays roles in skeletal muscle development and homeostasis. Moreover, its expression is upregulated in muscle undergoing atrophy as well as after muscle injury. Here we show that VDR regulates simulated microgravity-induced atrophy in C2C12 myotubes in vitro. After 8 h of microgravity simulated using 3D-clinorotation, the VDR-binding motif was associated with chromatin regions closed by the simulated microgravity and enhancer regions inactivated by it, which suggests VDR mediates repression of enhancers. In addition, VDR was induced and translocated into the nuclei in response to simulated microgravity. VDR-deficient C2C12 myotubes showed resistance to simulated microgravity-induced atrophy and reduced induction of FBXO32, an atrophy-associated ubiquitin ligase. These results demonstrate that VDR contributes to the regulation of simulated microgravity-induced atrophy at least in part by controlling expression of atrophy-related genes., (© 2022. The Author(s).)

Exposure to microgravity affects astronauts' health in adverse ways. However, less is known about the extent to which fibroblast differentiation during the wound healing process is affected by the lack of gravity. One of the key steps of this process is the differentiation of fibroblasts into myofibroblasts, which contribute functionally through extracellular matrix production and remodeling. In this work, we utilized collagen-based three-dimensional (3D) matrices to mimic interstitial tissue and studied fibroblast differentiation under simulated microgravity (sµG). Our results demonstrated that alpha-smooth muscle actin (αSMA) expression and translocation of Smad2/3 into the cell nucleus were reduced upon exposure to sµG compared to the 1 g control, which suggests the impairment of fibroblast differentiation under sµG. Moreover, matrix remodeling and production were decreased under sµG, which is in line with the impaired fibroblast differentiation. We further investigated changes on a transcriptomic level using RNA sequencing. The results demonstrated that sµG has less effect on fibroblast transcriptomes, while sµG triggers changes in the transcriptome of myofibroblasts. Several genes and biological pathways found through transcriptome analysis have previously been reported to impair fibroblast differentiation. Overall, our data indicated that fibroblast differentiation, as well as matrix production and remodeling, are impaired in 3D culture under sµG conditions.

Spaceflight associated neuro-ocular syndrome (SANS) is hypothesized to develop as a consequence of the chronic headward fluid shift that occurs in sustained weightlessness. We exposed healthy subjects (n = 24) to strict 6° head-down tilt bed rest (HDTBR), an analog of weightlessness that generates a sustained headward fluid shift, and we monitored for ocular changes similar to findings that develop in SANS. Two-thirds of the subjects received a daily 30-min exposure to artificial gravity (AG, 1 g at center of mass, ~0.3 g at eye level) during HDTBR by either continuous (cAG, n = 8) or intermittent (iAG, n = 8) short-arm centrifugation to investigate whether this intervention would attenuate headward fluid shift-induced ocular changes. Optical coherence tomography images were acquired to quantify changes in peripapillary total retinal thickness (TRT), retinal nerve fiber layer thickness, and choroidal thickness, and to detect chorioretinal folds. Intraocular pressure (IOP), optical biometry, and standard automated perimetry data were collected. TRT increased by 35.9 µm (95% CI, 19.9-51.9 µm, p < 0.0001), 36.5 µm (95% CI, 4.7-68.2 µm, p = 0.01), and 27.6 µm (95% CI, 8.8-46.3 µm, p = 0.0005) at HDTBR day 58 in the control, cAG, and iAG groups, respectively. Chorioretinal folds developed in six subjects across the groups, despite small increases in IOP. Visual function outcomes did not change. These findings validate strict HDTBR without elevated ambient CO 2 as a model for investigating SANS and suggest that a fluid shift reversal of longer duration and/or greater magnitude at the eye may be required to prevent or mitigate SANS., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society. This article has been contributed to by US Government employees and their work is in the public domain in the USA.)

All terrestrial organisms have evolved and adapted to thrive under Earth's gravitational force. Due to the increase of crewed space flights in recent years, it is vital to understand how the lack of gravitational forces affects organisms. It is known that astronauts who have been exposed to microgravity suffer from an array of pathological conditions including an impaired immune system, which is one of the most negatively affected by microgravity. However, at the cellular level a gap in knowledge exists, limiting our ability to understand immune impairment in space. This review highlights the most significant work done over the past 10 years detailing the effects of microgravity on cellular aspects of the immune system.

The blood-brain barrier (BBB) is critical to maintaining central nervous system (CNS) homeostasis. However, the effects of microgravity (MG) on the BBB remain unclear. This study aimed to investigate the influence of simulated MG (SMG) on the BBB and explore its potential mechanism using a proteomic approach. Rats were tail-suspended to simulate MG for 21 days. SMG could disrupt the BBB, including increased oxidative stress levels, proinflammatory cytokine levels, and permeability, damaged BBB ultrastructure, and downregulated tight junctions (TJs) and adherens junctions (AJs) protein expression in the rat brain. A total of 554 differentially expressed proteins (DEPs) induced by SMG were determined based on the label-free quantitative proteomic strategy. The bioinformatics analysis suggested that DEPs were mainly enriched in regulating the cell-cell junction and cell-extracellular matrix biological pathways. The inhibited Ras-related C3 botulinum toxin substrate 1 (Rac1)/Wiskott-Aldrich syndrome protein family verprolin-homologous protein 2 (Wave2)/actin-related protein 3 (Arp3) pathway and the decreased ratio of filamentous actin (F-actin) to globular actin contributed to BBB dysfunction induced by SMG. In the human brain microvascular endothelial cell (HBMECs), SMG increased the oxidative stress levels and proinflammatory cytokine levels, promoted apoptosis, and arrested the cell cycle phase. Expression of TJs and AJs proteins were downregulated and the distribution of F-actin was altered in SMG-treated HBMECs. The key role of the Rac1/Wave2/Arp3 pathway in BBB dysfunction was confirmed in HBMECs with a specific Rac1 agonist. This study demonstrated that SMG induced BBB dysfunction and revealed that Rac1/Wave2/Arp3 could be a potential signaling pathway responsible for BBB disruption under SMG. These results might shed a novel light on maintaining astronaut CNS homeostasis during space travel.

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 ˙ O 2max ) and maximum heart rate (HR max ) 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 ˙ O 2max 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 ˙ O 2 in space with implications for crew health and the physical deterioration of astronauts., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

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.

The human body experiences physiological changes under microgravity environment that phenocopy aging on Earth. These changes include early onset osteoporosis, skeletal muscle atrophy, cardiac dysfunction, and immunosenescence, and such adaptations to the space environment may pose some risk to crewed missions to Mars. To investigate the effect of microgravity on aging, many model organisms have been used such as the nematode Caenorhabditis elegans, the fruit fly Drosophila melanogaster, and mice. Herein we report that the budding yeast Saccharomyces cerevisiae show decreased replicative lifespan (RLS) under simulated microgravity in a clinostat. The reduction of yeast lifespan is not a result of decreased tolerance to heat shock or oxidative stress and could be overcome either by deletion of FOB1 or calorie restriction, two known interventions that extend yeast RLS. Deletion of the sirtuin gene SIR2 worsens the simulated microgravity effect on RLS, and together with the fob1Δ mutant phenotype, it suggests that simulated microgravity augments the formation of extrachromosomal rDNA circles, which accumulate in yeast during aging. We also show that the chronological lifespan in minimal medium was not changed when cells were grown in the clinostat. Our data suggest that the reduction in longevity due to simulated microgravity is conserved in yeast, worms, and flies, and these findings may have potential implications for future crewed missions in space, as well as the use of microgravity as a model for human aging., (Copyright © 2020 The Committee on Space Research (COSPAR). Published by Elsevier B.V. All rights reserved.)

Aboard the International Space Station (ISS), astronauts must adapt to altered vestibular and somatosensory inputs due to microgravity. Sensorimotor adaptation on Earth is often studied with a task that introduces visuomotor conflict. Retention of the adaptation process, known as savings, can be measured when subjects are exposed to the same adaptive task multiple times. It is unclear how adaptation demands found on the ISS might interfere with the ability to adapt to other sensory conflict at the same time. In the present study, we investigated the impact of 30 days' head-down tilt bed rest combined with elevated carbon dioxide (HDBR + CO 2 ) as a spaceflight analog on sensorimotor adaptation. Eleven subjects used a joystick to move a cursor to targets presented on a computer screen under veridical cursor feedback and 45° rotated feedback. During this NASA campaign, five individuals presented with optic disk edema, a sign of spaceflight-associated neuro-ocular syndrome (SANS). Thus, we also performed post hoc exploratory analyses between subgroups who did and did not show signs of SANS. HDBR + CO 2 had some impact on sensorimotor adaptation, with a lack of savings across the whole group. SANS individuals showed larger, more persistent after-effects, suggesting a shift from relying on cognitive to more implicit processing of adaptive behaviors. Overall, these findings suggest that HDBR + CO 2 alters the way in which individuals engage in sensorimotor processing. These findings have important implications for missions and mission training, which require individuals to adapt to altered sensory inputs over long periods in space. NEW & NOTEWORTHY This is the first bed rest campaign examining sensorimotor adaptation and savings in response to the combined effect of HDBR + CO2 and to observe signs of spaceflight-associated neuro-ocular syndrome (SANS) in HDBR participants. Our findings suggest that HDBR + CO2 alters the way that individuals engage in sensorimotor processing. Individuals who developed signs of SANS seem to rely more on implicit rather than cognitive processing of adaptive behaviors than subjects who did not present signs of SANS.

Deep-space missions may alter immune cell phenotype in the primary (e.g., thymus) and secondary (e.g., spleen) lymphoid organs contributing to the progression of a variety of diseases. In deep space missions, astronauts will be exposed to chronic low doses of HZE radiation while being in microgravity. Ground-based models of long-term uninterrupted exposures to HZE radiation are not yet available. To obtain insight in the effects of concurrent exposure to microgravity and chronic irradiation (CIR), mice received a cumulative dose of chronic 0.5 Gy gamma rays over one month ± simulated microgravity (SMG). To obtain insight in a dose rate effect, additional mice were exposed to single acute irradiation (AIR) at 0.5 Gy gamma rays. We measured proportions of immune cells relative to total number of live cells in the thymus and spleen, stress level markers in plasma, and change in body weight, food consumption, and water intake. CIR affected thymic CD3+/CD335+ natural killer T (NK-T) cells, CD25+ regulatory T (Treg) cells, CD27+/CD335- natural killer (NK1) cells and CD11c+/CD11b- dendritic cells (DCs) differently in mice subjected to SMG than in mice with normal loading. No such effects of CIR on SMG as compared to normal loading were observed in cell types from the spleen. Differences between CIR and AIR groups (both under normal loading) were found in thymic Treg and DCs. Food consumption, water intake, and body weight were less after coexposure than singular or no exposure. Compared to sham, all treatment groups exhibited elevated plasma levels of the stress marker catecholamines. These data suggest that microgravity and chronic irradiation may interact with each other to alter immune cell phenotypes in an organ-specific manner and appropriate strategies are required to reduce the health risk of crewmembers., (Copyright © 2020 The Committee on Space Research (COSPAR). Published by Elsevier B.V. All rights reserved.)

Long non-coding RNAs (lncRNAs) are important in regulating the response to environmental stresses in organisms. In this study, we used Caenorhabditis elegans as an animal model to determine the functions of intestinal lncRNAs in regulating response to simulated microgravity stress. Among the intestinal lncRNAs, linc-2, linc-46, linc-61, and linc-78 were increased by simulated microgravity treatment, and linc-13, linc-14, linc-50, and linc-125 were decreased by simulated microgravity treatment. Among these 8 intestinal lncRNAs, RNAi knockdown of linc-2 or linc-61 induced a susceptibility to toxicity of simulated microgravity, whereas RNAi knockdown of linc-13, linc-14, or linc-50 induced a resistance to toxicity of simulated microgravity. In simulated microgravity treated nematodes, linc-50 potentially binds to three transcriptional factors (DAF-16, SKN-1, and HLH-30). RNAi knockdown of daf-16, skn-1, or hlh-30 could suppress resistance of linc-50(RNAi) nematodes to the toxicity of simulated microgravity. Therefore, our results provide an important basis for intestinal lncRNAs, such as the linc-50, in regulating the response to simulated microgravity in nematodes.

Background: Circular RNAs (circRNAs) are a new class of non-coding RNA with a stable structure formed by special loop splicing. Research increasingly suggests that circRNAs play a vital role in the pathogenesis and progression of various diseases. However, the roles of circRNAs in osteoblast differentiation under microgravity remain largely unknown. Here, we investigated the roles and mechanobiological response of circRNAs in osteoblasts under simulated microgravity., Methods: Differential circRNA and mRNA expression profiles of MC3T3-E1 cells during exposure to microgravity were screened by RNA transcriptome sequencing technology (RNA-seq). The selected RNAs were validated using quantitative real-time polymerase chain reaction (qRT-PCR). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were applied for gene function analyses., Results: A total of 427 circRNAs and 1912 mRNAs were differentially expressed along with osteogenic differentiation in the simulated microgravity group (SMG) compared to the control group (CON). Of these, 232 circRNAs and 991 mRNAs were upregulated, whereas 95 circRNAs and 921 mRNAs were downregulated (fold change ≥ 2, p < 0.05). The results showed that the parental genes of circRNAs and mRNAs were mainly enriched in anatomical structure morphogenesis, anchoring junction and protein binding. KEGG analysis results showed that the differentially expressed mRNAs were enriched in the regulation of the actin cytoskeleton, focal adhesion, and Ras signalling pathway. Subsequently, 9 core regulatory genes, including 6 mRNAs and 3 circRNAs, were identified based on their possible function in osteoblast differentiation. Based on this analysis, circ&#95;014154 was selected as the target circRNA, which likely plays important roles in osteogenic differentiation processes under microgravity. The circRNA-miRNA-mRNA network showed that circRNAs might act as miRNA sponges to regulate osteoblast differentiation., Conclusion: By presenting a better understanding of the molecular mechanisms of genes and circRNAs in simulated microgravity, the present study will provide a novel view of circRNAs in the regulation of osteogenic differentiation and bone formation., (Copyright © 2020 Elsevier B.V. All rights reserved.)

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.

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., (© 2020 Federation of American Societies for Experimental Biology.)

Purpose: The objective of this study was to assess whether artificial gravity attenuates any long-duration head-down 6 0 bed rest (HDBR)-induced alterations in motor unit (MU) properties., Methods: Twenty-four healthy participants (16 men; 8 women; 26-54 years) underwent 60-day HDBR with (n = 16) or without (n = 8) 30 min artificial gravity daily induced by whole-body centrifugation. Compound muscle action potential (CMAP), MU number (MUNIX) and MU size (MUSIX) were estimated using the method of Motor Unit Number Index in the Abductor digiti minimi and tibialis anterior muscles 5 days before (BDC-5), and during day 4 (HDT4) and 59 (HDT59) of HDBR., Results: The CMAP, MUNIX, and MUSIX at baseline did not change significantly in either muscle, irrespective of the intervention (p > 0.05). Across groups, there were no significant differences in any variable during HDBR, compared to BDC-5., Conclusion: Sixty days of HDBR with or without artificial gravity does not induce alterations in motor unit number and size in the ADM or TA muscles in healthy individuals.

Dietary restriction has been well-described to improve health metrics, but whether it could benefit pathophysiological adaptation to extreme environment, for example, microgravity, remains unknown. Here, we investigated the effects of a daily rhythm of fasting and feeding without reducing caloric intake on cardiac function and metabolism against simulated microgravity. Male rats under ad libitum feeding or time-restricted feeding (TRF; food access limited to 8 hours every day) were subjected to hindlimb unloading (HU) to simulate microgravity. HU for 6 weeks led to left ventricular dyssynchrony and declined cardiac function. HU also lowered pyruvate dehydrogenase (PDH) activity and impaired glucose utilization in the heart. All these were largely preserved by TRF. TRF showed no effects on HU-induced loss of cardiac mass, but significantly improved contractile function of cardiomyocytes. Interestingly, TRF raised liver-derived fibroblast growth factor 21 (FGF21) level and enhanced cardiac FGF21 signaling as manifested by upregulation of FGF receptor-1 (FGFR1) expression and its downstream markers in HU rats. In isolated cardiomyocytes, FGF21 treatment improved PDH activity and glucose utilization, consequently enhancing cell contractile function. Finally, both liver-specific knockdown (KD) of FGF21 and cardiac-specific FGFR1 KD abrogated the cardioprotective effects of TRF in HU rats. These data demonstrate that TRF improves cardiac glucose utilization and ameliorates cardiac dysfunction induced by simulated microgravity, at least partially, through restoring cardiac FGF21 signaling, suggesting TRF as a potential countermeasure for cardioprotection in long-term spaceflight., (© 2020 Federation of American Societies for Experimental Biology.)

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., (© 2020 Federation of American Societies for Experimental Biology.)

The present study aimed to investigate the effect of microglia on simulated microgravity-induced hippocampal neurogenesis reduction and the possible mechanism underlying. Adult rats were treated with tail suspension for different times and the changes of neural stem cells (NSCs) were examined by immunohistochemistry. Then, minocycline was used to inhibit the activation of microglia, and the numbers of microglia and NSCs were detected after microgravity. Additionally, liquid protein chip analysis was applied to detect proinflammatory factors in hippocampus in order to find out the cytokines responsible for microglia activation after microgravity. The results revealed that microgravity increased the numbers of Iba1 + cells and decreased the numbers of BrdU + and DCX + cells in hippocampus but did not affect the ratio of NeuN + /BrdU + cells to the total number of BrdU + cells. After treated with minocycline, activated microglia were suppressed and the reduction of NSCs induced by microgravity recovered. Besides, compared with the control, higher concentrations of INF- γ and TNF- α were detected in the rats treated with microgravity. Our study provides the first evidence that microglia-mediated inflammation plays an important part in microgravity-induced neurogenesis reduction in hippocampus, and INF- γ and TNF- α secreted by microglia might be the key factors in this process., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Tian Lin et al.)

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., (© 2019 Wiley Periodicals, Inc.)

Regarding future space exploration missions and long-term exposure experiments, a detailed investigation of all factors present in the outer space environment and their effects on organisms of all life kingdoms is advantageous. Influenced by the multiple factors of outer space, the extremophilic bacterium Deinococcus radiodurans has been long-termly exposed outside the International Space Station in frames of the Tanpopo orbital mission. The study presented here aims to elucidate molecular key components in D. radiodurans, which are responsible for recognition and adaptation to simulated microgravity. D. radiodurans cultures were grown for two days on plates in a fast-rotating 2-D clinostat to minimize sedimentation, thus simulating reduced gravity conditions. Subsequently, metabolites and proteins were extracted and measured with mass spectrometry-based techniques. Our results emphasize the importance of certain signal transducer proteins, which showed higher abundances in cells grown under reduced gravity. These proteins activate a cellular signal cascade, which leads to differences in gene expressions. Proteins involved in stress response, repair mechanisms and proteins connected to the extracellular milieu and the cell envelope showed an increased abundance under simulated microgravity. Focusing on the expression of these proteins might present a strategy of cells to adapt to microgravity conditions.

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.

Intestinal epithelial cell (IEC) junctions constitute a robust barrier to invasion by viruses, bacteria and exposure to ingested agents. Previous studies showed that microgravity compromises the human immune system and increases enteropathogen virulence. However, the effects of microgravity on epithelial barrier function are poorly understood. The aims of this study were to identify if simulated microgravity alters intestinal epithelial barrier function (permeability), and susceptibility to barrier-disrupting agents. IECs (HT-29.cl19a) were cultured on microcarrier beads in simulated microgravity using a rotating wall vessel (RWV) for 18 days prior to seeding on semipermeable supports to measure ion flux (transepithelial electrical resistance (TER)) and FITC-dextran (FD4) permeability over 14 days. RWV cells showed delayed apical junction localization of the tight junction proteins, occludin and ZO-1. The alcohol metabolite, acetaldehyde, significantly decreased TER and reduced junctional ZO-1 localization, while increasing FD4 permeability in RWV cells compared with static, motion and flask control cells. In conclusion, simulated microgravity induced an underlying and sustained susceptibility to epithelial barrier disruption upon removal from the microgravity environment. This has implications for gastrointestinal homeostasis of astronauts in space, as well as their capability to withstand the effects of agents that compromise intestinal epithelial barrier function following return to Earth.

The overall objective of this study was to test the hypothesis that +Gz (hypergravity/positive acceleration) and microgravity can both aggravate intervertebral disc degeneration (IVDD). Due to +Gz and microgravity, many pilots develop IVDD. However, the lack of animal models of IVDD under conditions of simulated +Gz and microgravity has hampered research on the onset and prevention of IVDD. Rabbits were randomly allotted to a control group, microgravity group, +Gz group, or mixed (+Gz + microgravity) group. A tail-suspension model was utilized to simulate a microgravity environment and an animal centrifuge to mimic +Gz conditions. After exposure to the above conditions for 4, 8, and 24 weeks, the body weights (BW) of animals in the control group gradually increased over time, while those of animals in the microgravity and mixed groups both decreased (p < 0.001). As compared with the control group, the proteoglycan content of animals in the other three groups was significantly reduced (F = 192.83, p < 0.001). The imageological, histopathological, and immunohistochemical changes to the L6-S1 intervertebral disc samples suggests that the effects of +Gz and microgravity can aggravate IVDD over time. The mixed effects of +Gz and microgravity had the greatest effect on degeneration and +Gz had a particularly greater effect than microgravity.

Caenorhabditis elegans is useful for assessing biological effects of spaceflight and simulated microgravity. The molecular response of organisms to simulated microgravity is still largely unclear. Mitochondrial unfolded protein response (mt UPR) mediates a protective response against toxicity from environmental exposure in nematodes. Using HSP-6 and HSP-60 as markers of mt UPR, we observed a significant activation of mt UPR in simulated microgravity exposed nematodes. The increase in HSP-6 and HSP-60 expression mediated a protective response against toxicity of simulated microgravity. In simulated microgravity treated nematodes, mitochondria-localized ATP-binding cassette protein HAF-1 and homeodomain-containing transcriptional factor DVE-1 regulated the mt UPR activation. In the intestine, a signaling cascade of HAF-1/DVE-1-HSP-6/60 was required for control of toxicity of simulated microgravity. Therefore, our data suggested the important role of mt UPR activation against the toxicity of simulated microgravity in organisms.

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.

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.

Gravitational stress in general and microgravity (µg) in particular are regarded as major stress factors responsible for immune system dysfunction in space. To assess the effects of alternating µg and hypergravity (hyper-g) on immune cells, the attachment of peripheral blood mononuclear cells (PBMCs) to adhesion molecules under flow conditions and the antigen-induced immune activation in whole blood were investigated in parabolic flight (PF). In contrast to hyper-g (1.8 g) and control conditions (1 g), flow and rolling speed of PBMCs were moderately accelerated during µg-periods which were accompanied by a clear reduction in rolling rate. Whole blood analyses revealed a "primed" state of monocytes after PF with potentiated antigen-induced pro-inflammatory cytokine responses. At the same time, concentrations of anti-inflammatory cytokines were increased and monocytes displayed a surface molecule pattern that indicated immunosuppression. The results suggest an immunologic counterbalance to avoid disproportionate immune responses. Understanding the interrelation of immune system impairing and enhancing effects under different gravitational conditions may support the design of countermeasures to mitigate immune deficiencies in space.

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.

Rodent ground-based models are widely used to understand the physiological consequences of space flight on the physiological system and have been routinely employed since 1979 and the development of hind limb unloading (HLU). However, the next steps in space exploration now include to travel to Mars where the gravity is 38% of Earth's gravity. Since no human being has experienced this level of partial gravity, a sustainable ground-based model is necessary to investigate how the body, already impaired by the time spent in microgravity, would react to this partial load. Here, we used our innovative partial weight-bearing (PWB) model to mimic a short mission and stay on Mars to assess the physiological impairments in the hind limb muscles induced by two different levels of reduced gravity applied in sequential fashion. This could provide a safe, ground-based model to study the musculoskeletal adaptations to gravitational change and to establish effective countermeasures to preserve astronauts' health and function.

During space flight, microgravity has several negative effects on cognitive functions and learning and memory abilities. However, there are few effectively preventive methods that have been developed yet. Previous studies showed that repetitive transcranial magnetic stimulation (rTMS), as a novel non-invasive technique, alleviated cognitive dysfunctions and facilitated synaptic plasticity. In the present study, we used a hindlimb unloading (Hu) mouse model to simulate microgravity conditions. And then, we investigated whether rTMS played a neuroprotective role in a Hu mouse model. Behavioral experiments including Open field test and Novel object recognition test were performed. These results showed that spontaneous activity and recognition memory were reduced by Hu, while rTMS significantly protected against the harmful effect. Furthermore, electrophysiological recordings were performed to examine the level of synaptic plasticity including paired-pulse facilitation (PPF) and long-term potentiation (LTP). In the hippocampus DG and CA1 regions, dendritic spine density was measured using Golgi-Cox staining. Our data showed that rTMS effectively impeded the impairment of PPF and LTP, as well as the decrease of spine density induced by Hu. Subsequently, Western blot assay showed that rTMS inhibited the downregulation of CREB/BDNF signaling network associated proteins in Hu mice. It suggests that rTMS pre-treatment plays a neuroprotective role in protecting against cognitive impairments and synaptic plasticity deficits induced by microgravity stimulation., (Copyright © 2018 Elsevier B.V. All rights reserved.)

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., (© 2018 Wiley Periodicals, Inc.)

Space radiation and microgravity (μ G ) 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 μ G 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 μ G 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 µ G. Recently we incorporated a μ G ⁻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 μ G 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 μ G 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.

Most studies utilizing fish to study the effects of simulated microgravity (SMG) only observe the effects during the first week of development. They also do not take into account the potential impact on development of vibrations caused by the equipment. In this study we analyze the effects of both SMG and vibration on development of the skeleton. We analyze three different exposure durations and starting points that coincide with cranial neural crest cell migration. We use a combination of bone staining and morphometrics to analyze the effects. Our data show that both vibration and SMG affect vertebra number and body size; however, not all vertebrae are equally affected by each treatment. We also show that delayed ossification manifests during development, particularly after SMG exposure, and this translates into buckled and bent bones in adults. This study highlights the large impact of even very short exposure periods when they coincide with critical time points of development.

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., Competing Interests: Salvatore Rinaldi and Vania Fontani invented REAC technology. Giulia Cugia is affiliated with the commercial organization Virostatic s.r.l., Alghero, Italy. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Head-down tilt bed rest (HDBR) has been used as a spaceflight analog to study some of the effects of microgravity on human physiology, cognition, and sensorimotor functions. Previous studies have reported declines in balance control and functional mobility after spaceflight and HDBR. In this study we investigated how the brain activation for foot movement changed with HDBR. Eighteen healthy men participated in the current HDBR study. They were in a 6° head-down tilt position continuously for 70 days. Functional MRI scans were acquired to estimate brain activation for foot movement before, during, and after HDBR. Another 11 healthy men who did not undergo HDBR participated as control subjects and were scanned at four time points. In the HDBR subjects, the cerebellum, fusiform gyrus, hippocampus, and middle occipital gyrus exhibited HDBR-related increases in activation for foot tapping, whereas no HDBR-associated activation decreases were found. For the control subjects, activation for foot tapping decreased across sessions in a couple of cerebellar regions, whereas no activation increase with session was found. Furthermore, we observed that less HDBR-related decline in functional mobility and balance control was associated with greater pre-to-post HDBR increases in brain activation for foot movement in several cerebral and cerebellar regions. Our results suggest that more neural control is needed for foot movement as a result of HDBR. NEW & NOTEWORTHY Long-duration head-down bed rest serves as a spaceflight analog research environment. We show that brain activity in the cerebellum and visual areas during foot movement increases from pre- to post-bed rest and then shows subsequent recovery. Greater increases were seen for individuals who exhibited less decline in functional mobility and balance control, suggestive of adaptive changes in neural control with long-duration bed rest.

The present study aimed to determine how the learning and memory gradually change with the prolonged hindlimb unloading (HU) treatment in rats. Different HU durations (7 d, 14 d, 21 d, and 28 d) in Sprague-Dawley (SD) rats were implemented. Cognitive function was assessed using the Morris water maze (MWM) and the shuttle box test. Additionally, parameters about cholinergic activity and oxidative stress were tested. Results showed that longer-than-14 d HU led to the inferior performances in the behavioral tasks. Besides, acetylcholine esterase (AChE) activity, malondialdehyde (MDA) level in brain, reactive oxygen species (ROS), and 8-hydroxy-2-deoxyguanosine (8-OHdG) concentrations of HU rats were significantly increased. Furthermore, choline acetyltransferase (ChAT), superoxide dismutase (SOD), and catalase (CAT) activity in brain were notably attenuated. Most of these effects were more pronounced after longer exposure (21 d and 28 d) to HU, although some indicators had their own characteristics of change. These results indicate that cholinergic dysfunction and oxidative damage were involved in the learning and memory impairments induced by longer-than-14 d HU. Moreover, the negative effects of HU tend to be augmented as the HU duration becomes longer. The results may be helpful to present possible biochemical targets for countermeasures development regarding the memory deficits under extreme environmental conditions.

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., (Copyright © 2018 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.)

Microgravity has been previously demonstrated to induce skeletal muscle atrophy, loss of muscle force and disorders in myogenesis and metabolism. Current pharmacological strategies exhibit poor efficacy. Bu Zhong Yi Qi decoction (BZ) is a well‑known traditional Chinese medicine decoction used for myasthenia gravis. In the present study, its effect on unloading induced muscle atrophy was investigated. The mousetail suspension model was used to simulate weightlessness induced muscle atrophy. The results indicated that BZ could significantly protect muscles from simulated weightlessness‑induced atrophy. To elucidate the underlying mechanisms, drugCIPHER‑CS methods were introduced to predict its potential targets, significantly enriched pathways and biological processes. The results demonstrated that the calcium signaling pathway, citrate cycle, biosynthetic and lipid metabolic process are affected by BZ. Among the targets, nuclear receptor corepressor 1 (NCoR1) is one of the most important proteins involved in myogenesis and metabolism. The results indicated that BZ significantly downregulated NCoR 1 expression, and further induced muscle differentiation and metabolism by regulating NCoR1‑associated gene expression in vivo and in vitro. In summary, the present study indicated that may be effective in combating weightlessness‑induced muscle atrophy. Combined with bioinformatics, the underlying mechanism for this decoction was investigated, which provided an improved understanding of this decoction.

Background: Adverse effects of spaceflight on sensorimotor function have been linked to altered somatosensory and vestibular inputs in the microgravity environment. Whether these spaceflight sequelae have a central nervous system component is unknown. However, experimental studies have shown spaceflight-induced brain structural changes in rodents' sensorimotor brain regions. Understanding the neural correlates of spaceflight-related motor performance changes is important to ultimately develop tailored countermeasures that ensure mission success and astronauts' health., Method: Head down-tilt bed rest (HDBR) can serve as a microgravity analog because it mimics body unloading and headward fluid shifts of microgravity. We conducted a 70-day 6° HDBR study with 18 right-handed males to investigate how microgravity affects focal gray matter (GM) brain volume. MRI data were collected at 7 time points before, during and post-HDBR. Standing balance and functional mobility were measured pre and post-HDBR. The same metrics were obtained at 4 time points over ~90 days from 12 control subjects, serving as reference data., Results: HDBR resulted in widespread increases GM in posterior parietal regions and decreases in frontal areas; recovery was not yet complete by 12 days post-HDBR. Additionally, HDBR led to balance and locomotor performance declines. Increases in a cluster comprising the precuneus, precentral and postcentral gyrus GM correlated with less deterioration or even improvement in standing balance. This association did not survive Bonferroni correction and should therefore be interpreted with caution. No brain or behavior changes were observed in control subjects., Conclusions: Our results parallel the sensorimotor deficits that astronauts experience post-flight. The widespread GM changes could reflect fluid redistribution. Additionally, the association between focal GM increase and balance changes suggests that HDBR also may result in neuroplastic adaptation. Future studies are warranted to determine causality and underlying mechanisms.

Astronauts aboard the International Space Station (ISS) have exhibited hyperopic shifts, posterior eye globe flattening, dilated optic nerve sheaths, and even optic disk swelling from spaceflight. Elevated intracranial pressure (ICP) consequent to cephalad fluid shifts is commonly hypothesized as contributing to these ocular changes. Head-down tilt (HDT) is frequently utilized as an Earth-based analog to study similar fluid shifts. Sealed environments like the ISS also exhibit elevated CO 2 , a potent arteriolar vasodilator that could further affect cerebral blood volume (CBV) and cerebral blood flow, intracranial compliance, and ICP. A collaborative pilot study between the National Space Biomedical Research Institute and the German Aerospace Center tested the hypotheses that 1 ) HDT and elevated CO 2 physiologically interact and 2 ) cerebrovascular pulsatility is related to HDT and/or elevated CO 2 In a double-blind crossover study ( n = 6), we measured CBV pulsatility via near-infrared spectroscopy, alongside noninvasive ICP and intraocular pressure (IOP) during 28-h -12° HDT at both nominal (0.04%) and elevated (0.5%) ambient CO 2 In our cohort, CBV pulsatility increased significantly over time at cardiac frequencies (0.031 ± 0.009 μM/h increase in total hemoglobin concentration pulsatility amplitude) and Mayer wave frequencies (0.019 ± 0.005 μM/h increase). The HDT-CO 2 interaction on pulsatility was not robust but rather driven by one individual. Significant differences between atmospheres were not detected in ICP or IOP. Further work is needed to determine whether individual differences in pulsatility responses to CO 2 relate to visual changes in space. NEW & NOTEWORTHY Cerebral blood volume (CBV) pulsatility-as measured by near-infrared spectroscopy-increases over time during -12° head-down tilt at both cardiac and Mayer wave frequencies. CBV pulsatility appeared to increase more under elevated (0.5%) CO 2 at Mayer wave frequencies in some individuals. If similar dynamic pulsatility increases occur in astronauts, there is the potential to initiate vascular and possibly other remodeling processes that lead to symptoms associated with sustained increases in intracranial pressure.

Long-term spaceflight affects numerous organ systems in the body, including metabolic dysfunction. Recently, ample evidence has demonstrated that the liver is a vulnerable organ during spaceflight. However, the changes in hepatocyte proliferation and cell cycle control under microgravity remain largely unexplored. In the present study, we first confirmed that the serum levels of aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase, biochemical markers of liver function, were altered in rats under tail suspension (TS) conditions to simulate microgravity, as shown in previous reports. Next, we demonstrated that the cell proliferation activity, determined by Ki67, PCNA and PH3, was significantly decreased at the different TS time points (TS for 14, 28 and 42 days) compared with that in the control group. Consistently, the positive cell cycle regulators Ccna2, Ccnd1, Cdk1, Cdk2 and cyclin D3 were also significantly decreased in the TS groups as shown by quantitative real-time PCR and western blotting analysis. Subsequent analysis revealed that the aberrant hepatocyte proliferation inhibition under simulated microgravity was associated with the upregulation of miR-223 in the liver. We further found that miR-223 inhibited the proliferation of Hepa1-6 cells and identified CDK2 and CUL1 as its direct targets. In addition, the decreased expression of CDK2 and CUL1 was negatively correlated with the level of p27 in vitro and in vivo, which may have been responsible for retarding hepatocyte proliferation. Collectively, these data indicate that upregulation of miR-223 was associated with the inhibition of liver cell growth and reveal the role of miR-223 in rat hepatocyte proliferation disorders and the pathophysiological process under simulated microgravity.

Background: No studies have been published on an astronaut population to assess the effectiveness of countermeasures for limiting physiological changes in the lumbopelvic region caused by microgravity exposure during spaceflight. However, several studies in this area have been done using spaceflight simulation via bed-rest. The purpose of this systematic review was to evaluate the effectiveness of countermeasures designed to limit physiological changes to the lumbopelvic region caused by spaceflight simulation by means of bed-rest., Methods: Electronic databases were searched from the start of their records to November 2014. Studies were assessed with PEDro, Cochrane Risk of Bias and a bed-rest study quality tool. Magnitude based inferences were used to assess countermeasure effectiveness., Results: Seven studies were included. There was a lack of consistency across studies in reporting of outcome measures. Some countermeasures were found to be successful in preventing some lumbopelvic musculoskeletal changes, but not others. For example, resistive vibration exercise prevented muscle changes, but showed the potential to worsen loss of lumbar lordosis and intervertebral disc height., Conclusion: Future studies investigating countermeasures should report consistent outcomes, and also use an actual microgravity environment. Additional research with patient reported quality of life and functional outcome measures is advocated., (Copyright © 2016 Elsevier Ltd. All rights reserved.)