Project description:Genome-wide transcriptional profiling shows that reducing gravity levels in the International Space Station (ISS) causes important alterations in Drosophila gene expression. However, simulation experiments on ground, without space constraints, show weaker effects than space environment. A global and integrative analysis using the M-bM-^@M-^\gene expression dynamics inspectorM-bM-^@M-^] (GEDI) self-organizing maps, reveals a subtle response of the transcriptome using different populations and microgravity and hypergravity simulation devices. These results suggest that, in addition to behavioural responses that can be detected also at the gene expression level, the transcriptome is finely tuned to normal gravity. The alteration of this constant parameter on Earth can have effects on gene expression that depends both on the environmental conditions and the ground based facility used to compensate the gravity vector. Alternative and commons effects of mechanical facilities, like the Random Positioning Machine and a centrifuge, and strong magnetic field ones, like a cryogenically cooled superconductive magnet, are discussed. We compare the effects over the gene expression profile of different gender/age Drosophila imagoes in 3-4 days-long experiments under altered gravity conditions into three GBF ("Ground Based Facilities" for micro/hyper- gravity simulation) using whole genome microarray platforms. Descriptions of different GBFs ("treatments"): LDC means "Large Diameter Centrifuge". Samples can be placed under three conditions: inside LDC (at certain g level), at the LDC rotational control and at external 1g control (outside the LDC). RPM means "Random Positioning Machine". Samples can be placed under two conditions: inside RPM (at nearly 0g, Microgravity level) and at external 1g control (outside the RPM). At the magnet, means INSIDE the Magnetic levitator (another GBF). Samples can be placed under four conditions: inside Magnet 0g* (at microgravity with magnetic field), inside Magnet at 1g* (internal control with magnetic field) or inside the magnet 2g* (at hypergravity with magnetic field) and at external 1g control (outside the magnet)
Project description:au13-11_gravity - gravity - Cell cycle and cell proliferation are decoupled under altered gravity conditions. We have previously shown that semisolid cell cultures of Arabidopsis suffer overall genome changes in response to altered gravity and also that cell cycle stages duration is altered. By using synchronized cell cultures we will demonstrate the precise alterations in cell cycle duration and also the transcriptional signature in any of them. - Experiments consists on exposures of Arabidopsis cell cultures to 1g control/simulated microgravity (RPM) conditions. Asynchronous cells exposed for 14 h + Syncronous populations choosen to have an enrichment of cell cycle phases were used (being T7/T10 samples on G2 phase, T14/T16 samples on G1 phase).
Project description:au13-11_gravity - gravity - Cell cycle and cell proliferation are decoupled under altered gravity conditions. We have previously shown that semisolid cell cultures of Arabidopsis suffer overall genome changes in response to altered gravity and also that cell cycle stages duration is altered. By using synchronized cell cultures we will demonstrate the precise alterations in cell cycle duration and also the transcriptional signature in any of them. - Experiments consists on exposures of Arabidopsis cell cultures to 1g control/simulated microgravity (RPM) conditions. Asynchronous cells exposed for 14 h + Syncronous populations choosen to have an enrichment of cell cycle phases were used (being T7/T10 samples on G2 phase, T14/T16 samples on G1 phase). 6 dye-swap - time course,treated vs untreated comparison
Project description:The general objective of the study was to determine modulation of gene expression by environmental factors, with a special emphasis on bone formation. For this reason, the specific period of treatment was chosen between 5-6 days post-fertilization (dpf), when bone formation and calcification are taking place. This experiment was designed as a new type of gravitational experiment, which we like to call \relative microgravity\, referring to the fact that the larvae first grow in hyper gravity for 5 days and are then returned to 1g normal gravity for 1 day. Zebrafish embryos were placed on a Large Diameter Centrifuge at 3 hpf, brought to a gravitational force of 3 g until 5 dpf. Reference embryos were kept in parallel at 1g (Inc). At 5dpf, one batch was left at 3g (3g), one batch was returned to 1g (3g>1g), while a third batch was returned to 1g, but left on the axis of the centrifuge (Axe; 3g>Axe). The experiment was repeated 4 times, each time with 4 batches of 60 larvae.
Project description:SmallRNAs are proposed as key regulators in many cellular processes including angiogenesis, suggested candidates for future therapeutic applications. But, regulation and modulation of smallRNAs in pathology conditions and normal conditions are poorly recognized, supremely in wound care management. Current study focused to identify the smallRNA regulatory network in simulated microgravity sensitized Human Umbilical Cord Vein Endothelial cells (HUVECs) and gravity (1g) as a background. HUVECs were purchased from Lonza (Cat.No) and cultured in EGM2 medium (Cat.No CC3162) supplemented with 10% Fetal Bovine, 1% penicillin-streptomycin (W/V). Cells were cultured in RPM (Randomized positioned machine) for 2 hours at 37°C.
Project description:Efficient generation of functional cardiomyocytes from human induced pluripotent stem cells (hiPSC-CMs) is critical for their use in regenerative medicine and other applications. In this study, we evaluated the effect of space microgravity (µg) on the differentiation of hiPSC-derived cardiac progenitors compared with parallel 1g condition on the International Space Station. Cryopreserved 3D cardiac progenitors derived from hiPSCs were cultured for 3 weeks. Compared with 1g culture, the µg culture had larger sphere sizes, increased expression of proliferation markers, higher counts of nuclei, and higher cell viability. Highly enriched cardiomyocytes generated in µg had appropriate gene expression and cardiac structure as well as improved function including contractility and Ca2+ handling. RNA-seq analysis of 3-day cultures revealed that short-term exposure of cardiac progenitor spheres to space microgravity upregulated genes involved in cell proliferation, cardiac differentiation, and contraction. These results indicate that space microgravity increased survival and proliferation of hiPSC-CMs and improved their structures and functions.
Project description:Prolonged skeletal unloading through bedrest results in bone loss similar to that observed in elderly osteoporotic patients, but with an accelerated timeframe. This rapid effect on weight-bearing bones is also observed in astronauts who lose up to 2% of their bone mass per month spent in Space. Despite important implications for Spaceflight travellers and bedridden patients on Earth, the exact mechanisms involved in disuse osteoporosis have not been elucidated. Parathyroid hormone-related protein (PTHrP) regulates many physiological processes including skeletal development, and has been proposed as a gravisensor. To investigate the role of PTHrP in microgravity-induced bone loss, trabecular osteoblasts (TOs) from Pthrp+/+ and -/- mice were exposed to simulated microgravity for 6 days. Viability of TOs decreased in inverse proportion to PTHrP expression levels. Microarray analysis of Pthrp+/+ TOs after 6 days at 0g revealed expression changes in genes encoding prolactins,apoptosis and survival molecules, bone metabolism and extra-cellular matrix composition proteins, chemokines, IGF family and Wnt-related signalling molecules. Importantly, 88% of 0g-induced expression changes in Pthrp+/+ cells overlap those observed in Pthrp-/- cells in normal gravity. Pulsatile treatment with PTHrP1-36 peptide during microgravity exposure reversed a large proportion of 0g-induced changes in Pthrp+/+ TOs. Our results confirm PTHrP efficacy as an anabolic agent to prevent microgravity-induced cell death in TOs. Total RNA samples extracted from Pthrp+/+and -/- trabecular osteoblasts (TOs) exposed for 6 days to simulated 0g in Synthecon rotating cell, or left 6 days in culture at 1g. Cells had either been treated with a pulsatile treatment (2 h/day) of PTHrP1-36 peptide (10-8M) or received a change in growth medium. In total: 8 different conditions with 2 replicates each, i.e. Pthrp+/+ TOs at 0g or 1g with or without PTHrP1-36 treatment, and Pthrp-/- TOs at 0g or 1 g,with or without PTHrP1-36 treatment.
Project description:We developed a fast-rotating clinostat to simulate micrigravity (µg) and investigated various effects (including proliferation, self-renewal, and cell cycle regulation) of simulated microgravity (sµg) on human pluripotent stem cells (hPSC). Cells were cultured in sµg and control condition in normal gravity (1g). We observed significant upregulation of protein translation of human pluripotency transcription factors in hPSC cultured in sµg condition compared to 1g. We also noted a significant increase in the expression levels of genes involved in telomere elongation. Our induced differentiation experiments showed that hPSC cultured in sµg condition were less susceptible towards differentiation compared to cells cultured in 1g condition as indicated by the significant delayed in the process of differentiation of the cell in sµg condition. These results suggest that sµg conditions enhance the self-renewal of hPSC. Our study further revealed that sµg enhanced the cell proliferation of hPSC by regulating the expression of cell cycle associated kinases. Moreover, RNAseq analysis indicated that in sµg condition the expression of pathways related to differentiation and development and down-regulated, while multiple components of the ubiquitin proteasome system are up-regulated, thus further contributing to an enhanced self-renewal of hPSC. These effects of sµg were not replicated in human fibroblasts. Taken together, these results highlight pathways and mechanisms in hPSC vulnerable to µg that impose significant impacts on human health and performance, physiology, and cellular and molecular processes.
Project description:Jurkat T cells have been fixated at different gravity conditions during the 4th Swiss Parabolic Flight Campaign (4SPFC). 1g inflight samples were generated by crosslinking 5 minutes prior to onset of the first parabola. Hypg samples were generated by crosslinking the samples at the end of the first hypergravity 1.8 g phase, 20 seconds after start of the parabola. µg/0g samples were generated by crosslinking the cells 20 seconds after the onset of microgravity during the first parabola. A ground control reference condition was generated by crosslinking cells that were stored inside the flight hardware but were not onboard during the flight.