The interaction of microgravity and ethylene on the ultrastructure cell and Ca2+ localization in soybean hook hypocotyl

Calcium ions are secondary messenger in numerous cellular processes of plant grown at 1 g. Ca2+ are connected with oxygen atoms, of pectin carboxy groups and/or with H(+)-groups of protein (Roux and Slocum, 1982; Hepler and Wayne, 1985). The influence of altered gravity on the calcium balance in some cells is established. The increased synthesis of ethylene in plant grown in microgravity caused the change of the structural-functional organization of cell (Hensel and Iversen, 1980; Hilaire et al., 1996). Available data put the new question: how do high ethylene level and microgravity influence on the redistribution of Ca2+ in cell of seedling in early stage of growth? Therefore, the goal of our data was the comparable study of the cell ulltrastructure and localization of Ca2+ in hook hypocotyl of soybean seedling under interaction of microgravity and ethylene.     

微重力环境影响植物生长发育的研究进展

微重力是最独特的空间环境条件之一,研究微重力对不同植物种类以及不同植物部位的影响是空间生物学的重要内容之一,对于建立生物再生式生命保障系统意义重大。生物再生式生命保障系统是未来开展长期载人空间活动的核心技术,其优势在于能在一个密闭的系统内持续再生氧气,水和食物等高等动物生活必需品,植物部件是生物再生式生命保障系统的重要组成部分。了解和掌握微重力对植物生长发育的影响,有助于采取有效的作业制度确保其正常生长发育和繁殖,是成功建立生物再生式生命保障系统的首要关键。该文就植物在空间探索中的地位和作用,地面模拟微重力的装置以及国内外有关微重力对植物的影响做一综述。现有的研究结果包括,未来长期的载人航天任务需要植物通过光合作用为生物再生式生命保障系统提供部分动物营养、洁净水以及清除系统中的固体废物和二氧化碳;三维随机回旋装置是目前地面上模拟微重力效应的主要装置之一,尤其适用于植物材料的长期模拟微重力处理;国内外有关微重力对植物影响的报道生理生化水平多集中在植物的生长发育和生理反应,比如表型变化或者与重力相关的激素或者钙离子的再分配,细胞或亚细胞水平主要有细胞壁、线粒体、叶绿体以及细胞骨架等,基因和蛋白质表达水平的研究对象主要为拟南芥。由于实验方法和材料之间的差异,微重力对不同植物或者植物不同部位在各个水平的影响效果并不一致,未来需要开展更多的相关研究工作。     

Effects of microgravity on cell cytoskeleton and embryogenesis

The aim of this review is to compile, summarize and discuss the effects of microgravity on embryos, cell structure and function that have been demonstrated from data obtained during experiments performed in space or in altered gravity induced by clinostats. In cells and tissues cellular structure and genetic expression may be changed in microgravity and this has a variety of effects on embryogenesis which include death of the embryo, failure of neural tube closure, or final deformities to the overall morphology of the newborn or hatchling. Many species and protocols have been used for microgravity space experiments making it difficult to compare results. Experiments on the ways in which embryonic development and cell interactions occur in microgravity could also be performed. Experiments that have been done with cells in microgravity show changes in morphology, cytoskeleton and function. Changes in cytoskeleton have been noted and studies on microtubules in gravity have shown that they are gravity sensitive. Further study of basic chemical reactions that occur in cells should be done to shed some light on the underling processes leading to the changes that are observed in cells and embryos in microgravity.     

Cell-wall architecture and lignin composition of wheat developed in a microgravity environment.

The microgravity environment encountered during space-flight has long been considered to affect plant growth and developmental processes, including cell wall biopolymer composition and content. As a prelude to studying how microgravity is perceived - and acted upon - by plants, it was first instructive to investigate what gross effects on plant growth and development occurred in microgravity. Thus, wheat seedlings were exposed to microgravity on board the space shuttle Discovery (STS-51) for a 10 day duration, and these specimens were compared with their counterparts grown on Earth under the same conditions (e.g. controls). First, the primary roots of the wheat that developed under both microgravity and 1 g on Earth were examined to assess the role of gravity on cellulose microfibril (CMF) organization and secondary wall thickening patterns. Using a quick freeze/deep etch technique, this revealed that the cell wall CMFs of the space-grown wheat maintained the same organization as their 1 g-grown counterparts. That is, in all instances, CMFs were randomly interwoven with each other in the outermost layers (farthest removed from the plasma membrane), and parallel to each other within the individual strata immediately adjacent to the plasma membranes. The CMF angle in the innermost stratum relative to the immediately adjacent stratum was ca 80 degrees in both the space and Earth-grown plants. Second, all plants grown in microgravity had roots that grew downwards into the agar; they did not display "wandering" and upward growth as previously reported by others. Third, the space-grown wheat also developed normal protoxylem and metaxylem vessel elements with secondary thickening patterns ranging from spiral to regular pit to reticulate thickenings. Fourthly, both the space- and Earth-grown plants were essentially of the same size and height, and their lignin analyses revealed no substantial differences in their amounts and composition regardless of the gravitational field experienced, i.e. for the purposes of this study, all plants were essentially identical. These results suggest that the microgravity environment itself at best only slightly affected either cell wall biopolymer synthesis or the deposition of CMFs, in contrast to previous assertions.     

The biophysical limitations in physiological transport and exchange in plants grown in microgravity

This paper describes how changes in the physical behavior of fluids and gases in microgravity can limit the physiological transport and exchange in higher plants. These types of effects are termed indirect effects of microgravity because they are not due to gravity interacting with the mass of the plant body itself. The impact of limiting gravity-dependent transport phenomena has been analyzed by the use of mathematical modeling to simulate and compare biophysical transport in the 1g and spaceflight environments. These data clearly show that the microgravity environment induces significant limitations on basic physiological and biochemical processes within the aerial and rootzone portions of the plant. Furthermore, this mathematical model provides a solid foundation for explaining the physiological effects that have been noted in past spaceflight experiments.     

Cultivation of fall armyworm ovary cells in simulated microgravity

Summary A methodology is presented to culture Fall Armyworm Ovary cells in simulated microgravity using a novel bioreactor developed by NASA, the High-Aspect Ratio Vessel. In this vessel, the growth and metabolic profile for these insect cells were profoundly different than those obtained in shaker-flask culture. Specifically, stationary phase in the NASA vessel was extended from 24 h to at least 7 d while cell concentration and viability remained in excess of 1 × 10⁷ viable cells/ml and 90%, respectively. Measurements of glucose utilization, lactate production, ammonia production, and pH change indicate that simulated microgravity had a twofold effect on cell metabolism. Fewer nutrients were consumed and fewer wastes were produced in stationary phase by as much as a factor of 4 over that achieved in shaker culture. Those nutrients that were consumed in the NASA vessel were directed along different metabolic pathways as evidenced by an extreme shift in glucose utilization from consumption to production in lag phase and a decrease in yield coefficients by one half in stationary phase. These changes reflect a reduction in hydrodynamic forces from over 1 dyne/cm² in shaker culture to under 0.5 dyne/cm² in the NASA vessel. These results suggest that cultivation of insect cells in simulated microgravity may reduce production costs of cell-derived biologicals by extending production time and reducing medium requirements.     

Modeled microgravity affects motility and cytoskeletal structures

The hypothesis to be tested is that reduced cell-cell interactions between T cells and monocytes are one of the reasons for the observed depression of the "in vitro" activation of human lymphocytes in microgravity. Locomotion is essential for cell-cell contacts. Lymphocytes in suspension are highly motile in microgravity, whereas no data are available so far on the motility of adherent monocytes. It can be argued that an impaired locomotion of monocytes and cytoskeletal changes, both linked to cell contacts, could be responsible for their reduced interaction with T lymphocytes. This study is aimed at revealing how locomotion as well as cytoskeletal structures of adherent monocytes are modified under modeled microgravity conditions using the Random Positioning Machine (RPM, Dutch-Space) as earth based model of spaceflight.     

微重力条件下细胞培养和组织工程研究进展

随着空间生命科学研究的发展,人们将细胞、组织培养技术与微重力环境相结合产生了组织工程研究的一个新领域——微重力组织工程。模拟微重力条件下细胞培养和组织构建研究表明,微重力环境有利于细胞的三维生长,形成具有功能的组织样结构,培养后的三维组织无论从形态上还是基因表达上都更接近于正常的机体组织。这种微重力对细胞的作用效应,将可能为未来组织工程和再生医学研究提供一条新途径。该文概述了近十年来国内外微重力组织工程相关研究的最新进展。     

Space radiation effects and microgravity

Humans in space are exposed both to space radiation and microgravity. The question whether radiation effects are modified by microgravity is an important aspect in risk estimation. No interaction is expected at the molecular level since the influence of gravity is much smaller than that of thermal motion. Influences might be expected, however, at the cellular and organ level. For example, changes in immune competence could modify the development of radiogenic cancers. There are no data so far in this area. The problem of whether intracellular repair of radiation-induced DNA lesions is changed under microgravity conditions was recently addressed in a number of space experiments. The results are reviewed; they show that repair processes are not modified by microgravity.     

Physiology in microgravity.

Studies of physiology in microgravity are remarkably recent, with almost all the data being obtained in the past 40 years. The first human spaceflight did not take place until 1961. Physiological measurements in connection with the early flights were crude, but, in the past 10 years, an enormous amount of new information has been obtained from experiments on Spacelab. The United States and Soviet/Russian programs have pursued different routes. The US has mainly concentrated on relatively short flights but with highly sophisticated equipment such as is available in Spacelab. In contrast, the Soviet/Russian program concentrated on first the Salyut and then the Mir space stations. These had the advantage of providing information about long-term exposure to microgravity, but the degree of sophistication of the measurements in space was less. It is hoped that the International Space Station will combine the best of both approaches. The most important physiological changes caused by microgravity include bone demineralization, skeletal muscle atrophy, vestibular problems causing space motion sickness, cardiovascular problems resulting in postflight orthostatic intolerance, and reductions in plasma volume and red cell mass. Pulmonary function is greatly altered but apparently not seriously impaired. Space exploration is a new frontier with long-term missions to the moon and Mars not far away. Understanding the physiological changes caused by long-duration microgravity remains a daunting challenge.     

Amyloid fibril formation is suppressed in microgravity

In the future, humans may live in space because of global pollution and weather fluctuations. In microgravity, convection does not occur, which may change the amyloidogenicity of proteins. However, the effect of gravity on amyloid fibril formation is unclear and remains to be elucidated. Here, we analyzed the effect of microgravity on amyloid fibril formation of amyloidogenic proteins including insulin, amyloid β₄₂ (Aβ₄₂), and transthyretin (TTR). We produced microgravity (10^?3 g) by using the gravity controller Gravite. Human insulin, Aβ₄₂, and human wild-type TTR (TTRwt) were incubated at pH 3.0, 7.0, and 3.5 at 37 °C, respectively, in 1 g on the ground or in microgravity. We measured amyloidogenicity via the thioflavin T (ThT) method and cell-based 1-fluoro-2,5-bis[(E)-3-carboxy-4-hydroxystyryl]benzene (FSB) assay. ThT fluorescence intensity and cell-based FSB assay results for human insulin samples were decreased in microgravity compared with results in 1 g. Aβ₄₂ samples did not differ in ThT fluorescence intensity in microgravity and in 1 g on the ground. However, in the cell-based FSB assay, the staining intensity was reduced in microgravity compared with that on 1 g. Human TTRwt tended to form fewer amyloid fibrils in ThT fluorescence intensity and cell-based FSB assays in microgravity than in 1 g. Human insulin and Aβ₄₂ showed decreased amyloid fibril formation in microgravity compared with that in 1 g. Human TTRwt tended to form fewer amyloid fibrils in microgravity. Our experiments suggest that the earth's gravity may be an accelerating factor for amyloid fibril formation.     

模拟失重对白假丝酵母菌增殖和毒性的影响

【背景】近年来研究发现,失重条件可对一些致病微生物的增殖和毒性产生影响,白假丝酵母菌(Candida albicans)是典型的条件性致病真菌,在太空环境和人体中普遍存在,研究失重条件下白假丝酵母菌的增殖和毒性意义重大。【目的】利用旋转细胞培养系统(Rotary cell culture system,RCCS)模拟失重环境对白假丝酵母菌进行连续传代培养,检测模拟失重环境对白假丝酵母菌增殖情况、毒性以及基因表达的变化。【方法】将白假丝酵母菌接种在旋转生物反应器(High aspect rotating vessel,HARV)中,利用旋转细胞培养系统连续传代培养14 d,然后对菌株进行增殖速率测定、不同pH条件下增殖能力测定、生物膜相对形成能力测定和细胞毒性和动物毒力测定;利用转录组测序技术找出差异表达基因,结合性状分析模拟失重可能对白假丝酵母菌增殖和毒力的影响。【结果】与对照组相比,模拟失重组白假丝酵母菌对数期提前,增殖速率加快,在适宜pH条件下的增殖能力普遍提高,但其生物膜形成能力相对减弱,对LoVo细胞和小鼠的毒性减弱;转录组测序发现,模拟失重组共有280个基因表达差异达1.5倍以上(P0.05),其中248个上调、32个下调。差异基因经基因功能注释(Gene ontology,GO)和京都基因及基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)富集分析发现,相关胞膜形成及细胞分裂基因表达上调,生物膜形成、细胞黏附及共生粘连宿主基因表达下调。【结论】模拟失重环境可引起白假丝酵母菌增殖和毒性水平发生变化,相关改变可为研究失重环境对微生物的影响提供参考。     

How and why does the proteome respond to microgravity?

For medical and biotechnological reasons, it is important to study mammalian cells, animals, bacteria and plants exposed to simulated and real microgravity. It is necessary to detect the cellular changes that cause the medical problems often observed in astronauts, cosmonauts or animals returning from prolonged space missions. In order for in vitro tissue engineering under microgravity conditions to succeed, the features of the cell that change need to be known. In this article, we summarize current knowledge about the effects of microgravity on the proteome in different cell types. Many studies suggest that the effects of microgravity on major cell functions depend on the responding cell type. Here, we discuss and speculate how and why the proteome responds to microgravity, focusing on proteomic discoveries and their future potential.     

Simulated microgravity decreases DNA repair capacity and induces DNA damage in human lymphocytes

The effect of simulated microgravity on DNA damage and apoptosis is still controversial. The objective of this study was to test whether simulated microgravity conditions affect the expression of genes for DNA repair and apoptosis. To achieve this objective, human lymphocyte cells were grown in a NASA‐developed rotating wall vessel (RWV) bioreactor that simulates microgravity. The same cell line was grown in parallel under normal gravitational conditions in culture flasks. The effect of microgravity on the expression of genes was measured by quantitative real‐time PCR while DNA damage was examined by comet assay. The result of this study revealed that exposure to simulated microgravity condition decreases the expression of DNA repair genes. Mismatch repair (MMR) class of DNA repair pathway were more susceptible to microgravity condition‐induced gene expression changes than base excision repair (BER) and nucleotide excision repair (NER) class of DNA repair genes. Downregulation of genes involved in cell proliferation (CyclinD1 and PCNA) and apoptosis (Bax) was also observed. Microgravity‐induced changes in the expression of some of these genes were further verified at the protein level by Western blot analysis. The findings of this study suggest that microgravity may induce alterations in the expression of these DNA repair genes resulting in accumulation of DNA damage. Reduced expression of cell‐cycle genes suggests that microgravity may cause a reduction in cell growth. Downregulation of pro‐apoptotic genes further suggests that extended exposure to microgravity may result in a reduction in the cells' ability to undergo apoptosis. Any resistance to apoptosis seen in cells with damaged DNA may eventually lead to malignant transformation of those cells. J. Cell. Biochem. 107: 723–731, 2009. © 2009 Wiley‐Liss, Inc.     

Changes in gene expression and signal transduction in microgravity

Studies from space flights over the past three decades have demonstrated that basic physiological changes occur in humans during space flight. These changes include cephalic fluid shifts, loss of fluid and electrolytes, loss of muscle mass, space motion sickness, anemia, reduced immune response, and loss of calcium and mineralized bone. The cause of most of these manifestations is not known and until recently, the general approach was to investigate general systemic changes, not basic cellular responses to microgravity. This laboratory has recently studied gene growth and activation of normal osteoblasts (MC3T3-El) during spaceflight. Osteoblast cells were grown on glass coverslips and loaded in the Biorack plunger boxes. The osteoblasts were launched in a serum deprived state, activated in microgravity and collected in microgravity. The osteoblasts were examined for changes in gene expression and signal transduction. Approximately one day after growth activation significant changes were observed in gene expression in 0-G flight samples. Immediate early growth genes/growth factors cox-2, c-myc, bcl2, TGF beta1, bFGF and PCNA showed a significant diminished mRNA induction in microgravity FCS activated cells when compared to ground and 1-G flight controls. Cox-1 was not detected in any of the samples. There were no significant differences in the expression of reference gene mRNA between the ground, 0-G and 1-G samples. The data suggest that quiescent osteoblasts are slower to enter the cell cycle in microgravity and that the lack of gravity itself may be a significant factor in bone loss in spaceflight. Preliminary data from our STS 76 flight experiment support our hypothesis that a basic biological response occurs at the tissue, cellular, and molecular level in 0-G. Here we examine ground-based and space flown data to help us understand the mechanism of bone loss in microgravity.     

Lymphocyte movements and interactions in microgravity

Cell-cell contacts and the formation of aggregates play an important role in the mitogen induced in-vitro activation of lymphocytes. The fact that the formation of cell aggregates is only slightly reduced in microgravity suggests that cells are moving and interacting also in space. Direct evidence was obtained for the first time in an experiment performed on a sounding rocket flight, where the movements and interactions of free-floating, non activated cells could be observed in real time in microgravity. In an experiment performed on the IML-2 mission in July 1994, the movements and interactions of human lymphocytes during activation with the mitogen Con A were studied in real time in microgravity.     

Impairment of antigen-specific cellular immune responses under simulated microgravity conditions

Summary Microgravity has been implicated to play a role in the observed immune dysfunction of astronauts and cosmonauts after either short-term or long-term space travel. These reports, together with studies describing increased levels of microorganisms in the space cabin environment suggest potential risk for in-flight incidences of infectious diseases. In order to understand the mechanism underlying these immune defects, it is important to have a ground-based model that would reliably mimic the effects of microgravity on antigen-specific immune function. We tested the utility of the rotating wall vessel (RWV) technology developed at NASA as a model system because in the RWV the culture medium and the cells rotate synchronously with the vessel, thereby creating simulated microgravity conditions. We compared the RWV to the conventional tissue culture flask (T-flask), for culturing immune precursor cells with cytotoxic T lymphocyte (CTL) activity against synthetic viral peptides. We observed a significant loss of antigen-specific CTL activity in RWV cultures, but not in those from the T-flask, irrespective of the peptide immunogen used for inducing the primary immune response in different mouse strains. Loss of CTL activity in RWV cultures coincided with a significant reduction in CD8⁺ cells as well as CD4⁺ cells and DEC205⁺ dendritic cells, suggesting adverse effects of RWV culturing on both the effector and accessory cells for the loss of antigen-specific CTL function. These results provide a strong parallel to the reported defects in cell-mediated immunity during space travel and strongly support the utility of the RWV technology as an effective ground-based model for identifying key steps in immune cell dysfunction related to microgravity.