| Over the50-year-old history of human space flights it has beendemonstrated that the human body suffers from several conditions when it issubject to microgravity: bone mineral density is lost, muscle atrophy andcardiovascular deconditioning occur, pulmonary function and fluid regulatingsystems are impaired, sensory and balance systems are disturbed. In particular,space flights affect the cardiovascular system by compromising cardiovascularperformances, causing cardiac dysrhythmias, cardiac atrophy, orthostaticintolerance, and inducing a reduced aerobic capacity.Endothelial cells (ECs) cover the entire inner surface of the blood vesselsand play a crucial role in maintaining the functional integrity of the vascularwall. Functional properties of the endothelium include an active control of thevarious components of homeostasis, vasculartone and permeability and medial smooth muscle cell growth. ECs are highly sensitive to microgravity conditionand the morphological and functional changes of them are believed to be at thebasis of weightlessness-induced cardiovascular changes.Indeed, a number of studies have shown gravity-dependent modulations ofECs proliferation and differentiation. Microgravity seems to affect cellularorganization of the cytoskeleton, nitric oxide (NO) synthesis, intracellularsignaling mechanisms, gene expression and has controversial effects onapoptosis. In addition, ECs are protagonists in angiogenesis, i.e. the branchingand sprouting of capillaries from pre-existing blood vessels, which is a tightlycontrolled event crucial in development, wound healing and vacular remodeling.Many reports have indicated that the proliferation, migration and morphologicaldifferentiation of ECs play critical roles in the process of angiogenesis. However,the influence of microgravity on the ability of ECs to foster angiogenesisremains to be explored in detail.In the present study, we used a2D-clinostat to simulate microgravity, andwe observed tube formation, migration and expression of endothelial nitricoxide synthase (eNOS) in human umbilical vein endothelial cells (HUVEC-C).Specific inhibitors of eNOS (N-nitro-L-arginine methyl ester hydrochloride,L-NAME) and phosphoinositide3-kinase (PI3K) were added to the culturemedium and microgravity-induced changes in the pathways that mediateangiogenesis were investigated. The most significant and novel of the findingsdiscussed in these experiments are as below:1. The promotions of angiogenesis and migration induced by simulatedmicrogravity in HUVEC-C are correlated with eNOS activity. After24h ofexposure to simulated microgravity, HUVEC-C tube formation and migrationwere significantly promoted in comparison with the control groups, which could be reversed by co-incubation with L-NAME.2. Simulated microgravity induces up-regulation of eNOS in HUVEC-C.Immunofluorescence assay, RT-PCR and western blot analysis demonstratedthat eNOS expression in the HUVEC-C was significantly increased aftersimulated microgravity treatment. Ultrastructure observation via transmissionelectron microscope showed that the number of caveolae organelles in themembrane of HUVEC-C to be significantly reduced, which was correlate withenhanced activity of eNOS.3. The up-regulation of eNOS in HUVEC-C induced by simulatedmicrogravity is mediated by means of PI3K-Akt pathway. Furtherinvestigation with western blot analysis showed that phosphorylation of eNOSand serine/threonine kinase (Akt) were both up-regulated after exposure tosimulated microgravity. Moreover, the specific inhibitor of PI3K not onlysignificantly down-regulated the expression of phosphorylated Akt, but alsodown-regulated the eNOS phosphorylation. This suggested that PI3K-Akt signalpathway participate in modulating the activity of eNOS. In conclusion,24h ofexposure to simulated microgravity promotes angiogenesis among HUVEC-Cand this process is mediated through the PI3K-Akt-eNOS signal pathway.To sum up, the present study involves speculation of the underlying signaltransduction mechanisms of angiogenesis in HUVEC-C after exposure tosimulated microgravity via clinostat device. Our results suggest that24h ofsimulated microgravity can promote angiogenesis, migration, and eNOSexpression in HUVEC-C. Further investigations have confirmed that theincreased levels angiogenesis after simulated microgravity in HUVEC-C aremediated via PI3K-Akt-eNOS signal pathway. In this way, our studies reveal apreviously unrecognized role of simulated microgravity in ECs dysfunction. We not only plan a more detailed analysis of how modeled weightlessness affectsthe angiogenic process in ECs but also suggest new and complementaryexplanations to microgravity-induced vascular remodeling. |
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