Human cells, when exposed to both real and simulated microgravity (s-µ
), form 3D tissue constructs mirroring in vivo architectures (e.g., cartilage, intima constructs, cancer spheroids and others). ...In this study, we exposed human foetal osteoblast (hFOB 1.19) cells to a Random Positioning Machine (RPM) for 7 days and 14 days, with the purpose of investigating the effects of s-µ
on biological processes and to engineer 3D bone constructs. RPM exposure of the hFOB 1.19 cells induces alterations in the cytoskeleton, cell adhesion, extra cellular matrix (ECM) and the 3D multicellular spheroid (MCS) formation. In addition, after 7 days, it influences the morphological appearance of these cells, as it forces adherent cells to detach from the surface and assemble into 3D structures. The RPM-exposed hFOB 1.19 cells exhibited a differential gene expression of the following genes: transforming growth factor beta 1 (
, bone morphogenic protein 2 (
), SRY-Box 9 (
), actin beta (
), beta tubulin (
), vimentin (
), laminin subunit alpha 1 (
), collagen type 1 alpha 1 (
), phosphoprotein 1 (
) and fibronectin 1 (
). RPM exposure also induced a significantly altered release of the cytokines and bone biomarkers sclerostin (SOST), osteocalcin (OC), osteoprotegerin (OPG), osteopontin (OPN), interleukin 1 beta (IL-1β) and tumour necrosis factor 1 alpha (TNF-1α). After the two-week RPM exposure, the spheroids presented a bone-specific morphology. In conclusion, culturing cells in s-µ
under gravitational unloading represents a novel technology for tissue-engineering of bone constructs and it can be used for investigating the mechanisms behind spaceflight-related bone loss as well as bone diseases such as osteonecrosis or bone injuries.
Bone loss severely threatens the heath of astronauts in long-term spaceflight and the effects of the developed countermeasures are limited. Emerging studies have revealed the positive role of osthole ...(OST) in induction of osteogenic differentiation and bone formation. In this study, the effects of OST on bone metabolism were investigated in osteoblasts and rats treated with random positioning machine and hindlimb unloading, respectively. The results showed that OST treatment promoted the cellular proliferation and elevated the expression levels of RUNX2 and BMP2 in osteoblasts under simulated microgravity. In addition, hindlimb unloading rats administered with OST (5 mg kg/d, i.g.) exhibited improved bone mass, bone strength, expression of bone formation markers (BALP and OCN), and decreased expression of bone resorption markers (CTX-1 and TRACP-5b) in serum compared with rats treated with placebo (0.9% saline). Notably, we found that miR-34c-5p expression level was significantly up-regulated when exposed to simulated microgravity both in vitro and in vivo, and suppression of miR-34c-5p by antagomiR in rats obviously rescued the damages of BMD and trabecular bone microstructures caused by simulated microgravity. Importantly, OST treatment reversed miR-34c-5p expression level in hindlimb unloading rats, and ectopic expression of miR-34c-5p by transfection of agomiR in osteoblasts treated with OST decreased the expression levels of bone formation genes COL1α1, RUNX2, and BMP2. Overall, these data suggest an undisclosed role for OST in regulating microgravity-induced bone loss via control of miR-34c-5p expression, which may help to countermeasure development to overcome bone loss in spaceflight.
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•Osthole protects from bone loss induced by simulated microgravity.•Simulated microgravity induces up-regulation of miR-34c-5p expression level.•Osthole ameliorates simulated microgravity-induced bone loss via down-regulation of miR-34c-5p level.
Microgravity prominently affected cardiovascular health, which was the gravity-dependent physical factor. Deep space exploration had been increasing in frequency, but heart function was susceptible ...to conspicuous damage and cardiac mass declined in weightlessness. Understanding of the etiology of cardiac atrophy exposed to microgravity currently remains limited. The 3'-untranslated region (UTR) of casein kinase-2 interacting protein-1 (
) was a pivotal mediator in pressure overload-induced cardiac remodeling. However, the role of
3'-UTR in the heart during microgravity was unknown. We analyzed
mRNA 3'-UTR and coding sequence (CDS) expression levels in ground-based analogs such as mice hindlimb unloading (HU) and rhesus monkey head-down bed rest model.
3'-UTR had transcribed levels in the opposite change trend with cognate CDS expression in the hearts. We then subjected wild-type (WT) mice and cardiac-specific
3'-UTR-overexpressing mice to hindlimb unloading for 28 days. Our results uncovered that
3'-UTR remarkably attenuated cardiac dysfunction and mass loss in simulated microgravity environments. Mechanistically,
3'-UTR inhibited lipid accumulation and elevated fatty acid oxidation-related gene expression in the hearts through targeting calcium/calmodulin-dependent kinase 2 (CaMKK2) and activation of the AMPK-PPARα-CPT1b signaling pathway. These findings demonstrated
3'-UTR was an important regulator in atrophic heart growth after simulated microgravity.
Due to the limited self-repair capacity of articular cartilage, the surgical restoration of defective cartilage remains a major clinical challenge. The cell-based approach, which is known as ...autologous chondrocyte transplantation (ACT), has limited success, presumably because the chondrocytes acquire a fibroblast-like phenotype in monolayer culture. This unwanted dedifferentiation process is typically addressed by using three-dimensional scaffolds, pellet culture, and/or the application of exogenous factors. Alternative mechanical unloading approaches are suggested to be beneficial in preserving the chondrocyte phenotype. In this study, we examined if the random positioning machine (RPM) could be used to expand chondrocytes in vitro such that they maintain their phenotype. Bovine chondrocytes were exposed to (a) eight days in static monolayer culture; (b) two days in static monolayer culture, followed by six days of RPM exposure; and, (c) eight days of RPM exposure. Furthermore, the experiment was also conducted with the application of 20 mM gadolinium, which is a nonspecific ion-channel blocker. The results revealed that the chondrocyte phenotype is preserved when chondrocytes go into suspension and aggregate to cell clusters. Exposure to RPM rotation alone does not preserve the chondrocyte phenotype. Interestingly, the gene expression (mRNA) of the mechanosensitive ion channel
decreased with progressing dedifferentiation. In contrast, the gene expression (mRNA) of the mechanosensitive ion channel
was reduced around fivefold to 10-fold in all of the conditions. The application of gadolinium had only a minor influence on the results. This and previous studies suggest that the chondrocyte phenotype is preserved if cells maintain a round morphology and that the ion channel
could play a key role in the dedifferentiation process.
Cardiac atrophy and reduced cardiac distensibility have been reported following space flight. Cardiac function is correspondingly regulated in response to changes in loading conditions. Panax ...quinquefolium saponin (PQS) improves ventricular remodeling after acute myocardial infarction by alleviating endoplasmic reticulum stress and Ca2+overload. However, whether PQS can ameliorate cardiac atrophy following exposure to simulated microgravity remains unknown.
To explore the protective role of PQS in cardiac remodeling under unloading conditions and its underlying mechanisms.
Hindlimb unloading (HU) model was used to simulate unloading induced cardiac remodeling. Forty-eight male rats were randomly assigned to four groups, including control, PQS, HU and HU + PQS. At 8 weeks after the experiment, cardiac structure and function, serum levels of Creatine Kinase-MB (CK-MB), Cardiactroponin T (cTnT), ischemia modified albumin (IMA), and cardiomyocyte apoptosis were measured. Network pharmacology analysis was used to predict the targets of the six major constituents of PQS, and the signaling pathways they involved in were analyzed by bioinformatics methods. Changes in the key proteins involved in the protective effects of PQS were further confirmed by Western Blot.
Simulated microgravity led to increases in serum levels of CK-MB, cTnT and IMA, remodeling of cardiac structure, impairment of cardiac function, and increased cardiomyocyte apoptosis as compared with control. PQS treatment significantly reduced serum levels of CK-MB, cTnT and IMA, improved the impaired cardiac structure and function, and decreased cardiomyocyte apoptosis induced by unloading. The activation of AMPK and inhibition of Erk1/2 and CaMKII/HDAC4 were demonstrated in the cardiocytes of HU rats after PQS treatment.
PQS provides protection against cardiac remodeling induced by simulated microgravity, partly resulting from changes in the signaling pathways related to energy metabolism reduction, calcium overloading and cell apoptosis.
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Human cardiovascular system has adapted to Earth's gravity of 1G. The microgravity during space flight can induce cardiac remodeling and decline of cardiac function. At present, the mechanism of ...cardiac remodeling induced by microgravity remains to be disclosed. Casein kinase-2 interacting protein-1 (CKIP-1) is an important inhibitor of pressure-overload induced cardiac remodeling by decreasing the phosphorylation level of HDAC4. However, the role of CKIP-1 in the cardiac remodeling induced by microgravity is unknown. The purpose of this study was to determine whether CKIP-1 was also involved in the regulation of cardiac remodeling induced by microgravity. We first detected the expression of CKIP-1 in the heart from mice and monkey after simulated microgravity using Q-PCR and western blotting. Then, myocardial specific CKIP-1 transgenic (TG) and wild type mice were hindlimb-suspended (HU) to simulate microgravity effect. We estimated the cardiac remodeling in morphology and function by histological analysis and echocardiography. Finally, we detected the phosphorylation of AMPK, ERK1/2, and HDAC4 in the heart from wild type and CKIP-1 transgenic mice after HU. The results revealed the reduced expression of CKIP-1 in the heart both from mice and monkey after simulated microgravity. Myocardial CKIP-1 overexpression protected from simulated microgravity-induced decline of cardiac function and loss of left ventricular mass. Histological analysis demonstrated CKIP-1 TG inhibited the decreases in the size of individual cardiomyocytes of mice after hindlimb unloading. CKIP-1 TG can inhibit the activation of HDAC4 and ERK1/2 and the inactivation of AMPK in heart of mice induced by simulated microgravity. These results demonstrated CKIP-1 was a suppressor of cardiac remodeling induced by simulated microgravity.
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.
Microgravity can cause body fluids to accumulate in the brain, resulting in brain damage. There are few studies that focus on the detection of electrophysiological signals in simulated microgravity ...rats, and the precise mechanisms are unknown. In this study, a new device was established to investigate the influence of microgravity on hippocampal neurons. A 16-channel microelectrode array was fabricated for in vivo multichannel electrophysiological recordings. In these experiments, microelectrode array was inserted into normal, 28-day tail suspension model, and 3-day recovered after modulation rats to record electrophysiological signals in the CA1 and DG regions of the hippocampus. Through analysis of electrophysiological signals, we obtained the following results: (1) spike signals of model rats sporadically showed brief periods of suspension involving most of the recorded neurons, which corresponded to slow and smooth peaks in local field potentials. For model rats, the firing rate was reduced, and the power in the frequency spectrum was concentrated in the slow frequency band (0–1 Hz); (2) after the detected hippocampal cells divided into pyramidal cells and interneurons, the spike duration of pyramidal cells showed remarkable latency, and their average firing rates showed a more significant decrease compared to interneurons. These results demonstrate that the hippocampal neurons were impaired after modulation in the cellular dimension, and pyramidal cells were more susceptible than interneurons.
This work provided a novel nano-modified 16-site implantable microelectrode array (MEA) biosensor used for in vivo test of simulated microgravity (SM) rats. In summary, the highlights of the work are:•It is the first time to investigate electrophysiological signals on SM rats.•The MEA was fabricated with fine structure sharing a similar size with nerve cell.•16 sites at different depths designed for simultaneous detection in hippocampus.
A long-term stay of humans in space causes health problems and changes in protists and plants. Deep space exploration will increase the time humans or rodents will spend in microgravity (µ
). ...Moreover, they are exposed to cosmic radiation, hypodynamia, and isolation. OMICS investigations will increase our knowledge of the underlying mechanisms of µ
-induced alterations
and
.
We summarize the findings over the recent 3 years on µ
-induced changes in the proteome of protists, plants, rodent, and human cells. Considering the thematic orientation of microgravity-related publications in that time frame, we focus on medicine-associated findings, such as the µ
-induced antibiotic resistance of bacteria, the myocardial consequences of µ
-induced calpain activation, and the role of MMP13 in osteoarthritis. All these point to the fact that µ
is an extreme stressor that could not be evolutionarily addressed on Earth.
In conclusion, when interpreting µ
-experiments, the direct, mostly unspecific stress response, must be distinguished from specific µ
-effects. For this reason, recent studies often do not consider single protein findings but place them in the context of protein-protein interactions. This enables an estimation of functional relationships, especially if these are supported by epigenetic and transcriptional data (multi-omics).
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