Adhesion, germination and hyphae elongation of/from spores of Aspergillus tonophilus on a silica aerogel were observed, to indicate the possibility of mold contamination of the material designed for astrobiology mission using Kibo module in ISS. Spore germination seemed to be encouraged by nose sebum treatment before adhesion. Spore adhesion was observed by electron microscopy, and autofluorescence from mycelia was observed both by conventional microscope and USB-probe type microscope. The use of latter device was indicated to be effective for the monitoring of possible mold contamination of aerogel samples returned to earth in the astrobiology mission.
Tuber formation in potato (Solanum tuberosum L.) under hypergravity conditions in a centrifuge and simulated microgravity (μG) conditions on a horizontal clinostat was studied using in vitro culture of single-nodal stem segments prepared from sprouting tuber shoots. Hypergravity conditions at 100 G for 6 days reduced the growth of axillary bud (or stolon) of a single-nodal stem segment; the length, diameter, and fresh weight of the stolon developed under hypergravity conditions were 53%, 66%, and 74% of 1 G control, respectively. Promotion of swelling seen in the shoot under hypergravity conditions was not observed in the stolon. The size of cells and starch grains in the epidermal tissues in the swelling stolon were reduced under hypergravity conditions. On the other hand, the length of the stolon and the size of the tuber developed in the single-nodal stem segment under simulated μG conditions on a horizontal clinostat for 3 weeks were significantly increased as compared with the 1 G control. These results suggest that stolon growth and tuber formation in the single-nodal stem segment are under the control of gravity. Further studies on the molecular mechanism how gravity regulates tuber development and starch synthesis will be required for increasing potato tuber production in space.
Soy glycinin contains a functional inhibitory sequence, DIpYNP, against muscle-atrophy-associated ubiquitin ligase Cbl-b. It inhibited the binding of Cbl-b and IRS-1 (called Cblin-like peptide) and improved denervation-induced muscle atrophy in mice. In the present study, we evaluated the anti-muscle atrophy effect by feeding Kori-tofu to a mouse model of muscle atrophy induced by sciatic nerve transection, because Kori-tofu is a freeze-dried tofu made mainly from soy proteins. In the mice fed with Kori-tofu for one week, no significant inhibition in the reduction of the tibialis anterior (TA), gastrocnemius (GA) or soleus (SOL) muscle due to denervation was observed, compared with those of mice fed a soy protein isolate (SPI) or casein. However, in the TA muscle of the Kori-tofu-fed group, a significant suppression of the increased expression levels of muscle atrophy-related genes, MAFbx/atrogin-1 and MuRF-1, induced by denervation was observed similarly as that of mice fed a SPI. Additionally, the denervation-mediated decrease in the fast-twitch type myosin heavy chain (MyHC) level was suppressed in the TA of the Kori-tofu-fed group. Thus, our results suggest that Kori-tofu could be a useful and perishable functional space food against unloading-mediated muscle atrophy.
3-Hydroxy-3-methylglutaryl-CoA reductase (HMGR) catalyzes the rate-limiting step of the mevalonate pathway for biosynthesis of isoprenoids, such as sterols. An Arabidopsis T-DNA insertion mutant for HMGR1 (hmg1) produced roots, hypocotyls, rosette leaves, and inflorescences after germination at on-orbit 1 g in the Resist Tubule space experiment. The processes up to rosette leaf growth were normal in the hmg1 mutant under microgravity conditions; however, bolting of the inflorescence stem was strongly suppressed. The expression of a series of genes involved in sterol biosynthesis, including sterol glycosyltransferases implicated in producing lipid raft component steryl glycosides, and some of the raft-associated genes in the apical region of inflorescences in the α-tubulin 4 mutant (tua4) was suppressed under microgravity conditions. In addition, the fluorescence due to the raft marker remorin (REM1.3) was uniformly distributed along the rim of the cells of Arabidopsis hypocotyls in microgravity, as in the on-orbit control, showing the presence of lipid rafts in the plasma membrane under microgravity conditions. These results suggest that a severe reduction in sterol levels due to the hmg1 mutation, in combination with downregulation of the expression of sterol biosynthesis genes, causes the suppression of bolting in Arabidopsis under microgravity conditions in space and that lipid rafts may be involved in the bolting.
DNA analysis was performed by PCR followed by agarose gel electrophoresis in order to examine the method’s effectiveness for the human-cell contamination of returned samples in an astrobiology project (the Tanpopo mission). The effect of aerogel fragments on cell-derived DNA detection was investigated by focusing on Alu elements—primate-specific short interspersed sequences. A human cell-contaminated model of aerogel was prepared, and as a result, ca. 240 cells attached to the hydrophobic aerogel fragments were detected. No false positive was measured, indicating that our method is useful for evaluating the human contamination of aerogel returned from the Tanpopo mission.
Human astronauts follow a fixed prebreathing protocol consisting of 100% hyperoxia exposure repeated several times weekly before conducting the extravehicular activities in interplanetary space. However, despite astronaut exposure to both space radiation and hyperoxia, which have different biological effects on cells, the combinatorial effects on aging and longevity is unknown. Here, we evaluated the interactive effects on aging using Caenorhabditis elegans exposed to short-term 60Co γ-ray irradiation and hyperoxia. Thus, combined treatment extended the lifespan by approximately 10-15% in wild-type worms compared with irradiation only. Moreover, we found that the interaction induced not only the expression of antioxidant genes via insulin/IGF-1 signaling and the reduction of mitochondrial ROS, but also the activation of DNA damage repair system and sirtuin homologue genes during aging. Despite short-term irradiation of low linear energy transfer (LET) radiation such as γ-ray inducing transiently transcriptional inactivation of most genes, the levels of genes expression were recovered under hyperoxia during aging, at least partially. These findings in the C. elegans, showing that genetic transcriptional and mitochondrial dynamics depend on the interaction of low LET radiation with hyperoxia to consequently boost longevity, suggest some intracellular molecular targets to protect astronauts in space.
Growth and development, and polar auxin transport in epicotyls of etiolated pea (Pisum sativum L. cv Alaska) seedlings grown under hypergravity conditions produced by centrifugation were studied. Seeds were allowed to germinate and grow under various magnitude of hypergravity conditions for 3.5 days in the dark. Hypergravity at up to 50 G had little effect on epicotyl elongation growth, but increasing gravity force up to 300 G resulted in a reduction of elongation growth, the reduction at 100 G and 200 G being ca. 30% and 50% of control, respectively. Although expressions of genes playing an important role in polar auxin transport, auxin efflux carrier PsPINs and influx carrier PsAUX1, were not affected by hypergravity conditions, polar auxin transport activities of the epicotyls were substantially reduced, those at 100 G and 200 G being ca. 80% and 60% of control, respectively. In addition, close relationships between inhibition of elongation growth and polar auxin transport of the epicotyls were observed. These results suggest that hypergravity-regulated polar auxin activity is involved in inhibition of elongation growth of epicotyls of etiolated pea seedlings. The mechanisms to regulate elongation growth and polar auxin transport are discussed.
Exposure to microgravity during space flight has caused astronauts to experience decreased bone density, resulting from bone resorption by osteoclastic activation. Activation of the nuclear factor-κB ligand (RANKL) of the RANKL-producing cells, a known osteoclastogenesis-promoting factor, seems to be induced under microgravity. However, the role of the RANKL-producing cells under microgravity has not yet been demonstrated due to the lack of suitable in vitro organ culture of osteoblasts, osteoclasts, and osteocytes with the intact bone matrix under microgravity. Previous reports demonstrated that RANKL-producing cells were detected with a specific antiserum for goldfish RANKL in the regenerating goldfish scales. In this study, the response of RANKL-producing cells to simulated microgravity with a three-dimensional clinostat was examined using in vitro organ culture system with regenerating scales. In addition, mRNA expression analysis of osteoclastic and osteoblastic markers via quantitative real-time PCR was conducted together with histological analysis of RANKL-producing cells. After 4 days of exposure to simulated microgravity, the number of RANKL immune-positive cells increased compared with the RANKL immune-positive cells in control regenerating scales. The Rankl mRNA expression in the regenerating scales after simulated microgravity treatments was significantly higher than that in the control (1G) scales. The mRNA expression of osteoprotegerin (Opg), an osteoclastogenesis inhibitory factor, significantly decreased under simulated microgravity conditions. The ratio of Rankl/Opg in the simulated microgravity-treated scales was significantly higher than that in the scales of 1G control. The change in the ratio of Rankl/Opg implied that osteoclastic activation is promoted after simulated microgravity treatments. Actually, in the regenerating scales exposed to simulated microgravity, the mRNA expression of osteoclastic markers, such as receptor activator of NFκB, cathepsin K, integrin beta-3, and cellular-Src, significantly increased. On the other hand, the mRNA expression of the osteoblastic markers (collagen type I alpha 1 and osteocalcin) significantly decreased. The changes in the osteoclastic and osteoblastic markers were consistent with our previously reported experiment on the International Space Station. To the best of our knowledge, we are the first to demonstrate the activation of RANKL-producing cells under simulated microgravity conditions by in vitro organ culture using regenerating goldfish scales. We strongly emphasize that fish scales serve as an excellent bone model for the analysis of gravitational responses while maintaining conditions similar to in vivo.
We conducted the Resist Tubule space experiment to clarify the role of the cell wall in the resistance of inflorescence stems of Arabidopsis (Arabidopsis thaliana) to gravitational acceleration in the Cell Biology Experiment Facility on the Kibo Module of the International Space Station. The cell wall rigidity of inflorescence stems was lower under microgravity conditions than at 1g, particularly in the basal region, which plays a central role in supporting the entire plant in the presence of gravity. In the basal region under microgravity conditions, the levels of matrix and cellulosic polysaccharides per unit length decreased in correlation with a decline in cell wall rigidity. The expression of cellulose synthase genes CESA4, CESA7, and CESA8, and certain xylan synthase genes tended toward suppression in the basal region under microgravity conditions. The downregulation of the expression of these genes may cause a decrease in cell wall polysaccharide levels, thereby maintaining a soft and extensile cell wall under microgravity conditions. In addition, the levels of UV-absorbing compounds decreased in pace with the cellulose levels in the basal region. Combined with the data of the hypergravity experiments, these results suggest that secondary walls in the basal supporting region play an important role in the resistance of Arabidopsis inflorescence stems to gravity in the range of 1 g to hypergravity.
Bones are important organs for body resistance against force produced by gravity, though the influence of gravity on bone development is unclear. To examine the effects of gravity on osteogenesis, medaka larvae were reared in water or gel under various conditions. For determining the effects on bone development in a state of motion, larvae were reared in water under normal gravity (1g) or hypergravity (5g) conditions. Also, to examine the direct effect of gravity on bone mineralization, larvae were embedded in low melting agarose gel containing alizarin complexone (ALC) and reared for three days under a normal gravity (1g), simulated-microgravity (s-μg) with use of a clinostat device, or 5g condition. Medaka reared in water under the 5g condition showed forward protruding jaws and spreading of the mineralized area of jaw teeth as compared to those reared under the 1g control condition. In addition, the direction of growth of the notochord in the fin region was changed upward in those reared under the 5g condition, accompanied by a part of acetylated tubulinpositive nerves also localized upward, while positive signals for DsRed, expressed by an osterix promoter, in osteoblasts were increased in the fin region. On the other hand, in medaka reared in gel, ALC signals in the fin ray of those in the s-μg condition were increased as compared to those in the 5g condition. Changes noted in medaka larvae over three days indicated osteogenesis adaptation to the specific gravity environment. The present results obtained with an experimental system are considered useful for examinations in the future regarding changes of osteogenesis, which will be needed to clarify the mechanism of the effects of gravity on bone development. ©2021 Jpn. Soc. Biol. Sci. Space; doi:10.2187/bss.35.24
To understand the combined effect of microgravity (μG) and space radiation on cells is an important requirement for the anticipated space travel by humans. Cultured cells, such as human fibroblasts and lymphoblasts, were used in previous studies, and we observed changes in the expression of some cell-cycle-related genes and increased chromosome aberrations (CAs) in these cells under simulated μG. In this study, we investigated the combined effect of μG and space radiation on human peripheral blood lymphocytes. Whole blood was irradiated with X-ray or carbon-ion (C-ion) beam while being exposed to simulated μG using a three- dimensional clinostat. The frequency of CA was assessed using the three-color fluorescence in situ hybridization on chromosome spreads of colcemid-induced prematurely condensed chromosomes in lymphocytes during the first cell division post irradiation. Compared with the cells irradiated at 1G, the frequency of CA was increased in cells simultaneously exposed to simulated μG and radiation even though cells were irradiated by the same doses (0.5–1.0 Gy). This result is similar to those of our previous studies in which we used human lymphoblast TK6 cells and human fibroblast 1BR-hTERT cells. These ground-based experiments give insights into the biological effects of the space environment where radiation and μG coexist.
Plant culture experiments in space require non-destructive and automated assessment of growth that is efficient and unattended. Currently, plant growth conditions in spacecraft are often captured by ordinary RGB (red, green blue) cameras. In addition to images in the RGB wavelengths, near-infrared (NIR) images that are not affected by plant pigments, such as chlorophylls, could be used to improve the quality of growth-related information. In this study, we examined the possibility of using NIR to assess the growth status of a model moss plant, Physcomitrium (Physcomitrella) patens. Mosses cultured in multi-well plates were imaged with a hyperspectral (HS) sensor, and the length of gametophores, the number of leaves and gametophores, and the amount of photosynthetic pigments in the moss colonies were measured. Then we used the R, G, B, and NIR (850 nm) bands from the obtained HS images to extract the moss colony areas and evaluate the amount of growth. Our results suggested that spectral indices using R and NIR bands were suitable for assessing the morphology of moss colonies and extracting the colony area. The extracted area can be used to estimate the amount of growth.
This paper introduces the use of microarray data technology with the Agilent Maize Oligo Microarray (Design ID 016047) to characterize global changes in the transcript abundance of etiolated Zea mays (cv. Golden Cross Bantam) seedlings grown under microgravity (μg) conditions on the International Space Station (ISS) compared with those grown under 1 g conditions on Earth. Gene array data were analyzed according to stringent criteria that restricted the scored genes for specific hybridization values at least two fold. Of the 32152 - 32616 transcripts detected, 1030 and 590 transcripts were significantly different in the coleoptiles and in the mesocotyls. Of the transcripts detected, 877 and 428 transcripts were found to increase under μg conditions in the coleoptiles and the mesocotyls, respectively. Venn diagram analysis showed that 154 transcripts commonly increased and 10 decreased under μg conditions irrespective of the organ difference. Of these, phytohormone-related genes were focused, indicating that some of them were responsive to gravity. These results support the commonly accepted idea that phytohormone-related genes play a significant role in regulating plant growth and development under different gravity conditions.
We have previously reported that microgravity promotes the activation of osteoclasts in cultured regenerating scales. This osteoclastic activation was induced by increased levels of receptor activator of the nuclear factor-κB ligand (RANKL). Therefore, we determined that RANKL is an important factor in evaluating osteoclastogenesis in bone tissue. However, the role of RANKL in fish scales is poorly understood. In the present study, we prepared antiserum against goldfish RANKL in rabbits and detected RANKL-producing cells in regenerating goldfish scales. Furthermore, we studied osteoclastic activation by the addition of RANKL to examine exogenous RANKL on osteoclastogenesis in regenerating goldfish scales. As a result, RANKL immune-positive cells were detected in grooves of regenerating scales. In addition, treating the regenerating scales with mammalian RANKL for 3 h significantly increased the expression of the nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), which is essential for osteoclast differentiation. After 6 h of incubation with RANKL, the expression of cathepsin K, a functional osteoclastic gene, significantly increased. Furthermore, the molecules for osteoclast multinucleation and differentiation significantly increased following treatment with mammalian RANKL. Therefore, in fish scales as well as mammalian bone, we concluded that RANKL plays an important role in osteoclastogenesis.
The present paper is a comprehensive report on the Auxin Transport space experiment: the analysis of gravity response and attitude control mechanisms of plants under microgravity conditions in space on the International Space Station. The Auxin Transport space experiment was conducted in 2016 and 2017 in the Japanese Experiment Module (JEM) on the International Space Station (ISS), with the principal objective being integrated analyses of the growth and development of etiolated pea (Pisum sativum L. cv Alaska) and maize (Zea mays L. cv Golden Cross Bantam) seedlings under true microgravity conditions in space relative to polar auxin transport. To clarify auxin dynamics at molecular levels, gene expression of PsPIN1 and ZmPIN1a mRNA, and their products detected by immunohistochemistry were also investigated. In addition, the use of microarray data technology with Medicago ( Medicago truncatula ) microarrays to characterize global changes in the transcript abundance of etiolated Alaska pea seedlings grown under microgravity conditions in comparison with those under artificial 1 G conditions on the International Space Station was reported here. Comprehensive analyses of endogenous plant hormones in etiolated pea and maize seedlings grown under microgravity conditions in space as well as on 1 G conditions on Earth have already performed in the space experiment.
It is well-reported that the morphological properties of skeletal muscles or muscle fibers, which are influenced by the level of protein synthesis and/or degradation, are regulated in response to mechanical load. However, the precise mechanism responsible for such phenomena is not fully understood yet. Changes of the distribution of satellite cells and/or myonuclei have been also noted in atrophied or hypertrophied skeletal muscle fibers, suggesting that the number and/or function of these parameters play essential roles in the regulation of morphological properties of muscle and muscle fibers. Thus, the roles of satellite cells and/or myonuclei in the regulation of morphological properties of anti-gravitational muscle, soleus and adductor longus, in response to the level of mechanical stress, with or without association of macrophage-related factors, were briefly reviewed. It was suggested that a regulatory network among macrophage, interleukin-6, heat shock transcription factor 1, and activation of transcription factor 3 may play a crucial role for the modulation of skeletal muscle mass and function, which are also influenced by activation of satellite cells and distribution of myonuclei.
