Search Results (22)

Spaceflight induces a wide array of effects on the human body, notably including pathological changes mediated by alterations in gravity. Abnormalities in the formation of primary cilia (ciliogenesis) can lead to cell cycle arrest and decreased epithelial cell proliferation, thereby delaying wound healing. To investigate the effect of microgravity on ciliogenesis in bronchial epithelial cells, we used a 3D clinostat to generate simulated microgravity (SMG) conditions. When BEAS-2B bronchial epithelial cells were exposed to SMG for 72 h, their proliferation was significantly reduced. The expression of Ki-67, which is not expressed in the G0 phase, decreased under SMG. Conversely, the expression of p27, which is expressed in the G0 and G1 phases, increased under SMG. These results suggest that SMG led to an increase in the number of cells in the quiescent phase. When the mRNA expressions of ARL13B (a marker of cilia assembly) and disassembly-related genes (Aurora A, NDE1, HDAC6, and DVL2) were evaluated, SMG upregulated ciliary assembly markers and downregulated disassembly markers. In addition, SMG increased the cilia length and number of ciliated cells. These findings suggest that SMG contributes to reduced cell proliferation through cell cycle arrest by disrupting normal ciliogenesis. Our findings indicate that SMG could delay lung injury by decreasing cell proliferation. Full article

The extreme space environment accelerates aging and compromises human health. NASA has named five main hazards in space, including gravity changes. However, the contribution of each factor to the overall impact on biomolecular and cellular processes is not always clear. We aimed to explore the effects of microgravity on the transcriptomes of healthy volunteers, with a focus on gene expression in p53 pathways. Ten healthy men were exposed to dry immersion simulated microgravity (DI-SMG) for three weeks and blood samples were collected at five timepoints before, during and after the course of DI-SMG. T cells were purified from the peripheral blood samples and total RNA was isolated and sequenced followed by a bioinformatics analysis of the volunteers’ global transcriptomes. A differential expression of p53 network genes was observed. The expression of 30 genes involved in the p53 gene network was affected during a 3-week course of DI-SMG including classic p53 downstream target genes involved in cellular senescence: GADD45, p21, PUMA, IGF1 and other target genes. For the first time, the p53-associated cell signaling pathways and gene networks in human T cells were reported to be affected in vivo by DI-SMG. It is evident that the relatively mild effects of simulated weightlessness on the human body are sufficient to activate these pathways. Identified transcriptomic changes point toward a potential molecular overlap with aging and cellular senescence. These findings could contribute to a broader research landscape that may lead to the discovery of a new class of drugs—MG-senolytics. Full article

Pancreatic ductal adenocarcinoma (PDAC) accounts for 90% of all pancreatic malignancies. Despite the remarkable improvement concerning treatment, late detection and resistance to clinically used chemotherapeutic agents remain major challenges. Trichostatin A (TSA), a histone deacetylase inhibitor, has been recognized as an effective therapeutic agent against PDAC by inhibiting proliferation, inducing apoptosis, and sensitizing PDAC cells to chemotherapeutic agents such as gemcitabine. Microgravity has become a useful tool in cancer research due to its effects on various cellular processes. This paper presents a deep molecular and proteomic analysis investigating cell growth, the modulation of cytokeratins, and proteins related to apoptosis, cellular metabolism, and protein synthesis after TSA treatment in simulated microgravity (SMG)-exposed PaCa44 3D cells. Our analysis concerns the effects of TSA treatment on cell proliferation: the impairment of the cell cycle with the downregulation of proteins involved in Cdc42 signaling and G1/G2- and G2/M-phase transitions. Thus, we observed modification of survival pathways and proteins related to autophagy and apoptosis. We also observed changes in proteins involved in the regulation of transcription and the repair of damaged DNA. TSA treatment promotes the downregulation of some markers involved in the maintenance of the potency of stem cells, while it upregulates proteins involved in the induction and modulation of the differentiation process. Our data suggest that TSA treatment restores the cell phenotype prior to simulated microgravity exposure, and exerts an intriguing activity on PDAC cells by reducing proliferation and inducing cell death via multiple pathways. Full article

Space flight has many adverse effects on the physiological functions of astronauts. Certain similarities have been observed in some physiological processes of rodents and astronauts in space, although there are also differences. These similarities make rodents helpful models for initial investigations into space-induced physiological changes. This study uses a 3D-Clinostat to simulate microgravity and explores the role of microgravity in space flight-induced liver and brain abnormalities by comparing changes in the gut microbiota, serum metabolites, and the function and physiological biochemistry of liver and brain tissues between the simulated microgravity (SMG) group mice and the wild type (WT) group mice. The study, based on hematoxylin-eosin (HE) staining, 16S sequencing technology, and non-targeted metabolomics analysis, shows that the gut tissue morphology of the SMG group mice is abnormal, and the structure of the gut microbiota and the serum metabolite profile are imbalanced. Furthermore, using PICRUST 2 technology, we have predicted the functions of the gut microbiota and serum metabolites, and the results indicate that the liver metabolism and functions (including lipid metabolism, amino acid metabolism, and sugar metabolism, etc.) of the SMG group mice are disrupted, and the brain tissue metabolism and functions (including neurotransmitters and hormone secretion, etc.) are abnormal, suggesting a close relationship between microgravity and liver metabolic dysfunction and brain dysfunction. Additionally, the high similarity in the structure of the gut microbiota and serum metabolite profile between the fecal microbiota transplant (FMT) group mice and the SMG group mice, and the physiological and biochemical differences in liver and brain tissues compared to the WT group mice, suggest that microgravity induces imbalances in the gut microbiota, which in turn triggers abnormalities in liver and brain metabolism and function. Finally, through MetaMapp analysis and Pearson correlation analysis, we found that valeric acid, a metabolite of gut microbiota, is more likely to be the key metabolite that relates to microgravity-induced gut microbiota abnormalities, disorders of amino acid and lipid metabolism, and further induced metabolic or functional disorders in the liver and brain. This study has significant practical application value for deepening the understanding of the adaptability of living organisms in the space environment. Full article

The metabolic poise, or balance, between glycolysis and fatty acid oxidation (FAO) has recently been found to play a critical role in osteogenic differentiation and homeostasis. While simulated microgravity (SMG) is known to impede osteoblast differentiation (OBD) by inhibiting the Wnt/β-catenin pathway, how it affects osteoblast metabolism in this context remains unclear. We previously analyzed the effect of SMG on the differentiation of pre-osteoblast MC3T3-E1 cells and found that it reduced focal adhesion kinase (FAK) activity. This, in turn, downregulated Wnt/β-catenin and two of its downstream targets critical for OBD bone morphogenic protein-2 (BMP2) and type-1 collagen (COL1) formation, leading to a reduction in alkaline phosphatase (ALP) activity and cell matrix mineralization. In this study, we further analyzed how SMG-induced alterations in energy metabolism contribute to the inhibition of OBD in MC3T3-E1 cells. Consistent with our earlier findings, we demonstrated that SMG inhibits OBD by downregulating the collective activity of FAK and the Wnt/β-catenin-BMP2-COL1 transcriptional pathway. Interestingly, we observed that SMG also reduces the abundance of sirtuin-1 (SIRT1), peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) and carnitine palmitoyl transferase-1α (CPT1A), which are all key metabolic factors regulating mitochondrial number and FAO capacity. Accordingly, we found that the mitochondrial content and FAO potential of MC3T3-E1 cells were lower upon exposure to SMG but were both rescued upon administration of the FAK activator cytotoxic necrotizing factor-1 (CNF1), thereby allowing cells to overcome SMG-induced inhibition of OBD. Taken together, our study indicates that the metabolic regulator SIRT1 may be a new target for reversing SMG-induced bone loss. Full article

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. Full article

The space environment can affect the function of all physiological systems, including the properties of cell membranes. Our goal in this study was to explore the effect of simulated microgravity (SMG) on the cellular uptake of small molecules based on reported microgravity-induced changes in membrane properties. SMG was applied to cultured cells using a random-positioning machine for up to three hours. We assessed the cellular accumulation of compounds representing substrates of uptake and efflux transporters, and of compounds not shown to be transported by membrane carriers. Exposure to SMG led to an increase of up to 60% (p < 0.01) in the cellular uptake of efflux transporter substrates, whereas a glucose transporter substrate showed a decrease of 20% (p < 0.05). The uptake of the cathepsin activity-based probe GB123 (MW, 1198 g/mol) was also enhanced (1.3-fold, p < 0.05). Cellular emission of molecules larger than ~3000 g/mol was reduced by up to 50% in SMG (p < 0.05). Our findings suggest that short-term exposure to SMG could differentially affect drug distribution across membranes. Longer exposure to microgravity, e.g., during spaceflight, may have distinct effects on the cellular uptake of small molecules. Full article

Background: Cells are sensitive to changes in gravity, especially the cytoskeletal structures that determine cell morphology. The aim of this study was to assess the effects of simulated microgravity (SMG) on 3T3 cell morphology, as demonstrated by a characterization of the morphology of cells and nuclei, alterations of microfilaments and microtubules, and changes in cycle progression. Methods: 3T3 cells underwent induced SMG for 72 h with Gravite^®, while the control group was under 1G. Fluorescent staining was applied to estimate the morphology of cells and nuclei and the cytoskeleton distribution of 3T3 cells. Cell cycle progression was assessed by using the cell cycle app of the Cytell microscope, and Western blot was conducted to determine the expression of the major structural proteins and main cell cycle regulators. Results: The results show that SMG led to decreased nuclear intensity, nuclear area, and nuclear shape and increased cell diameter in 3T3 cells. The 3T3 cells in the SMG group appeared to have a flat form and diminished microvillus formation, while cells in the control group displayed an apical shape and abundant microvilli. The 3T3 cells under SMG exhibited microtubule distribution surrounding the nucleus, compared to the perinuclear accumulation in control cells. Irregular forms of the contractile ring and polar spindle were observed in 3T3 cells under SMG. The changes in cytoskeleton structure were caused by alterations in the expression of major cytoskeletal proteins, including β-actin and α-tubulin 3. Moreover, SMG induced 3T3 cells into the arrest phase by reducing main cell cycle related genes, which also affected the formation of cytoskeleton structures such as microfilaments and microtubules. Conclusions: These results reveal that SMG generated morphological changes in 3T3 cells by remodeling the cytoskeleton structure and downregulating major structural proteins and cell cycle regulators. Full article

The impact of space radiation and microgravity on DNA damage responses has been discussed controversially, largely due to the variety of model systems engaged. Here, we performed side-by-side analyses of human hematopoietic stem/progenitor cells (HSPC) and peripheral blood lymphocytes (PBL) cultivated in a 2D clinostat to simulate microgravity before, during and after photon and particle irradiation. We demonstrate that simulated microgravity (SMG) accelerates the early phase of non-homologous end joining (NHEJ)-mediated repair of simple, X-ray-induced DNA double-strand breaks (DSBs) in PBL, while repair kinetics in HSPC remained unaltered. Repair acceleration was lost with increasing LET of ion exposures, which increases the complexity of DSBs, precluding NHEJ and requiring end resection for successful repair. Such cell-type specific effect of SMG on DSB repair was dependent on the NF-кB pathway pre-activated in PBL but not HSPC. Already under unperturbed growth conditions HSPC and PBL suffered from SMG-induced replication stress associated with accumulation of single-stranded DNA and DSBs, respectively. We conclude that in PBL, SMG-induced DSBs promote repair of radiation-induced damage in an adaptive-like response. HSPC feature SMG-induced single-stranded DNA and FANCD2 foci, i.e., markers of persistent replication stress and senescence that may contribute to a premature decline of the immune system in space. Full article

The drug efflux transporter permeability glycoprotein (P-gp) plays an important role in oral drug absorption and distribution. Under microgravity (MG), the changes in P-gp efflux function may alter the efficacy of oral drugs or lead to unexpected effects. Oral drugs are currently used to protect and treat multisystem physiological damage caused by MG; whether P-gp efflux function changes under MG remains unclear. This study aimed to investigate the alteration of P-gp efflux function, expression, and potential signaling pathway in rats and cells under different simulated MG (SMG) duration. The altered P-gp efflux function was verified by the in vivo intestinal perfusion and the brain distribution of P-gp substrate drugs. Results showed that the efflux function of P-gp was inhibited in the 7 and 21 day SMG-treated rat intestine and brain and 72 h SMG-treated human colon adenocarcinoma cells and human cerebral microvascular endothelial cells. P-gp protein and gene expression levels were continually down-regulated in rat intestine and up-regulated in rat brain by SMG. P-gp expression was regulated by the Wnt/β-catenin signaling pathway under SMG, verified by a pathway-specific agonist and inhibitor. The elevated intestinal absorption and brain distribution of acetaminophen levels also confirmed the inhibited P-gp efflux function in rat intestine and brain under SMG. This study revealed that SMG alters the efflux function of P-gp and regulates the Wnt/β-catenin signaling pathway in the intestine and the brain. These findings may be helpful in guiding the use of P-gp substrate drugs during spaceflight. Full article

Microgravity is a novel strategy that may serve as a complementary tool to develop future cancer therapies. In lung cancer, the influence of microgravity on cellular processes and the migratory capacity of cells is well addressed. However, its effect on the mechanisms that drive lung cancer progression remains in their infancy. In this study, 13 differentially expressed genes were shown to be associated with the prognosis of lung cancer under simulated microgravity (SMG). Using gene set enrichment analysis, these genes are enriched in humoral immunity pathways. In lieu, alveolar basal-epithelial (A549) cells were exposed to SMG via a 2D clinostat system in vitro. In addition to morphology change and decrease in proliferation rate, SMG reverted the epithelial-to-mesenchymal transition (EMT) phenotype of A549, a key mechanism in cancer progression. This was evidenced by increased epithelial E-cadherin expression and decreased mesenchymal N-cadherin expression, hence exhibiting a less metastatic state. Interestingly, we observed increased expression of FCGBP, BPIFB, F5, CST1, and CFB and their correlation to EMT under SMG, rendering them potential tumor suppressor biomarkers. Together, these findings reveal new opportunities to establish novel therapeutic strategies for lung cancer treatment. Full article

Space microgravity condition has great physiological influence on astronauts’ health. The interaction of endothelial cells, which control vascular permeability and immune responses, is sensitive to mechanical stress. However, whether microgravity has significant effects on the physiological function of the endothelium has not been investigated. In order to address such a question, a clinostat-based culture model with a HUVEC monolayer being inside the culture vessel under the simulated microgravity (SMG) was established. The transmittance of FITC-tagged dextran was used to estimate the change of integrity of the adherens junction of the HUVEC monolayer. Firstly, we found that the permeability of the HUVEC monolayer was largely increased after SMG treatment. To elucidate the mechanism of the increased permeability of the HUVEC monolayer under SMG, the levels of total expression and activated protein levels of Rap1 and Rap2 in HUVEC cells, which regulate the adherens junction of endothelial cells, were detected by WB and GST pull-down after SMG. As the activation of both Rap1 and Rap2 was significantly decreased under SMG, the expression of Rap1GEF1 (C3G) and Rap1GAP in HUVECs, which regulate the activation of them, was further determined. The results indicate that both C3G and Rap1GAP showed a time-dependent increase with the expression of Rap1GAP being dominant at 48 h after SMG. The down-regulation of the expression of junctional proteins, VE-cadherin and β-catenin, in HUVEC cells was also confirmed by WB and immunofluorescence after SMG. To clarify whether up-regulation of Rap1GAP is necessary for the increased permeability of the HUVEC monolayer after SMG, the expression of Rap1GAP was knocked down by Rap1GAP-shRNA, and the change of permeability of the HUVEC monolayer was detected. The results indicate that knock-down of Rap1GAP reduced SMG-induced leaking of the HUVEC monolayer in a time-dependent manner. In total, our results indicate that the Rap1GAP-Rap signal axis was necessary for the increased permeability of the HUVEC monolayer along with the down-regulation of junctional molecules including VE-cadherin and β-catenin. Full article

Microgravity is known to impact bone health, similar to mechanical unloading on Earth. In the absence of countermeasures, bone formation and mineral deposition are strongly inhibited in Space. There is an unmet need to identify nutritional countermeasures. Curcumin and carnosic acid are phytonutrients with anticancer, anti-inflammatory, and antioxidative effects and may exhibit osteogenic properties. Zinc is a trace element essential for bone formation. We hypothesized that these nutraceuticals could counteract the microgravity-induced inhibition of osteogenic differentiation and function. To test this hypothesis, we cultured 7F2 murine osteoblasts in simulated microgravity (SMG) in a Random Positioning Machine in the presence and absence of curcumin, carnosic acid, and zinc and evaluated cell proliferation, function, and differentiation. SMG enhanced cell proliferation in osteogenic medium. The nutraceuticals partially reversed the inhibitory effects of SMG on alkaline phosphatase (ALP) activity and did not alter the SMG-induced reduction in the expression of osteogenic marker genes in osteogenic medium, while they promoted osteoblast proliferation and ALP activity in the absence of traditional osteogenic media. We further observed a synergistic effect of the intermix of the phytonutrients on ALP activity. Intermixes of phytonutrients may serve as convenient and effective nutritional countermeasures against bone loss in space. Full article

Astronauts are always faced with serious health problems during prolonged spaceflights. Previous studies have shown that weightlessness significantly affects the physiological function of female astronauts, including a change in reproductive hormones and ovarian cells, such as granulosa and theca cells. However, the effects of microgravity on these cells have not been well characterized, especially in granulosa cells. This study aimed to investigate the effects of simulated microgravity (SMG) on the proliferation and morphology of porcine granulosa cells (pGCs). pGC proliferation from the SMG group was inhibited, demonstrated by the reduced O.D. value and cell density in the WST-1 assay and cell number counting. SMG-induced pGCs exhibited an increased ratio of cells in the G0/G1 phase and a decreased ratio of cells in the S and G2/M phase. Western blot analysis indicated a down-regulation of cyclin D1, cyclin-dependent kinase 4 (cdk4), and cyclin-dependent kinase 6 (cdk6), leading to the prevention of the G1-S transition and inducing the arrest phase. pGCs under the SMG condition showed an increase in nuclear area. This caused a reduction in nuclear shape value in pGCs under the SMG condition. SMG-induced pGCs exhibited different morphologies, including fibroblast-like shape, rhomboid shape, and pebble-like shape. These results revealed that SMG inhibited proliferation and induced morphological changes in pGCs. Full article

Studies have shown that bone marrow-derived mesenchymal stem cells (BMSCs) can differentiate into dermal fibroblasts to participate in skin-repairing. However, at present, little is known about how microgravity affects dermal fibroblastic differentiation of BMSCs in space. The aim of this study was to investigate the effect of simulated microgravity (SMG) on the differentiation of BMSCs into dermal fibroblasts and the related molecular mechanism. Here, using a 2D-clinostat device to simulate microgravity, we found that SMG inhibited the differentiation and suppressed the Wnt/β-catenin signaling and phosphorylation of extracellular regulated protein kinases 1/2 (ERK1/2). After upregulating the Wnt/β-catenin signaling with lithium chloride (LiCl) treatment, we found that the effect of the differentiation was restored. Moreover, the Wnt/β-catenin signaling was upregulated when phosphorylation of ERK1/2 was activated with tert-Butylhydroquinone (tBHQ) treatment. Taken together, our findings suggest that SMG inhibits dermal fibroblastic differentiation of BMSCs by suppressing ERK/β-catenin signaling pathway, inferring that ERK/β-catenin signaling pathway may act as a potential intervention target for repairing skin injury under microgravity conditions. Full article