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Effects of hypergravity environment on lignin formation in Arabidopsis.

Lignin and the secondary wall formation are essential for evolution of land plants. In this study, effects of hypergravity environment on the morphology of the secondary wall and the lignin content were examined in Arabidopsis thaliana. Xylem vessels showed intense staining with phloroglucinol-HCl and autofluorescence under UV light at the basal region of the flower stalk when seedlings grown for 3 days after hypergravity treatment for 24 hours. And, the flower stalk exposed to hypergravity showed slight increase in the lignin content. These results suggest that the lignin formation is positively regulated under hypergravity.

Arabidopsis↗

Effect of hypergravity stimulus on XTH gene expression in Arabidopsis thaliana.

Hypergravity stimulus suppresses plant shoot growth by making the cell wall rigid. Xyloglucan endotransglucosylase/hydrolase (XTH) is involved in determining the rigidity of cell walls. We demonstrated that hypergravity influenced the expression of some XTH genes in shoots of Arabidopsis thaliana L.; in response to hypergravity stimulus of 300 g, the expression of AtXTH22 was up-regulated, while that of AtXTH15 was down-regulated. The effect of hypergravity on the expression of these genes was nullified by lanthanum chloride at 0.1 mM, suggesting that the expression of these XTH genes in Arabidopsis is under the control of the mechanoreceptor.

Arabidopsis↗

Space cycle: a human-powered centrifuge that can be used for hypergravity resistance training.

INTRODUCTION: This study represents the first step toward testing the hypothesis that hypergravity can be used as a unique resistance training modality for maintaining the health and function of skeletal muscle in microgravity. The primary objectives of this study were to use a human-powered short-arm centrifuge for the following: 1) to determine whether subjects could perform squats under hypergravity conditions without developing motion sickness or illusory motion; 2) to measure foot forces while performing squats under hypergravity conditions; and 3) to determine the mechanical power required to produce 1.5, 2.0, 2.5, and 3.0 Gz (head to foot) at the feet. METHODS: Subjects (22 males and 19 females) performed 10 squats each at 1.5, 2.0, 2.5, and 3.0 Gz on a human-powered short-arm centrifuge, the space cycle. Foot forces during each squat were monitored using insole force sensors, and normalized to foot forces measured at 1 Gz (relative foot forces). RESULTS: Illusory motion was minimized using a visual focal point, and did not affect the ability of subjects to perform squats. The mean standing relative foot forces at 3.0 Gz were 2.3 and 2.4 for the male and female subjects, respectively. The work rate required to power the space cycle was a linear function of Gz, and is well within the aerobic scope of untrained individuals. DISCUSSION: The findings of this study demonstrate that hypergravity can be used as an effective modality for loading skeletal muscle and that subjects can perform squat resistance exercise without developing motion sickness or illusory motion.

Adaptation, Physiological↗

Induction of early response genes by hypergravity in cultured mouse osteoblastic cells (MC3T3-E1).

Hypergravity as low as 50g transiently stimulated cultured mouse osteoblastic cells (MC3T3-E1) to induce early response genes such as c-fos and egr-1, whereas expression of c-jun was marginally affected. The maximum induction of c-fos required more than 90g, but egr-1 induction became maximum below 50g. Staurosporin inhibited the induction of c-fos by hypergravity almost completely at a concentration of 0.1 microM, but it inhibited the induction of egr-1 only partially. In cells pretreated with 12-O-tetradecanoylphorbol 13-acetate, induction of c-fos by hypergravity was almost completely abolished, whereas that of egr-1 was not affected. Activity of protein kinase C seemed to be activated in cells centrifuged at 900g. These results indicate that hypergravity stimulates multiple signal transduction cascades that are connected with the expression of early response genes.

Animals↗

Influence of hypergravity on the development of monoaminergic systems in the rat spinal cord.

We have investigated in this study the influence of a moderate hypergravity (1.8 G) on the development of monoaminergic projections to the spinal cord in the rat. Pregnant dams and their offspring were submitted to hypergravity from day 11 of gestation to postnatal day 15. Some animals were sacrificed at birth, other at postnatal day 15 and other after 8 months of normal gravity. In newborn animals, a substantial delay of the development of monoaminergic projections to the spinal cord was evidenced. In 15 days and 8 months animals, the pattern of innervation appeared anarchic, with numerous dystrophic profiles, mainly of serotonergic system. Ultrastructural examination of serotonergic projections revealed a paucity of synapses, and the frequent enveloping of serotonergic boutons by thin astrocytic profiles. We conclude that rats submitted to hypergravity during the critical period of onset of monoaminergic projections to the spinal cord are affected durably in the organization and the ultrastructure of these projections. Future studies are directed to the functional analysis of hypergravity animals, and to the influence of microgravity on the same system.

Animals↗

A mild stress due to hypergravity exposure at young age increases longevity in Drosophila melanogaster males.

Drosophila melanogaster flies were exposed to hypergravity starting at two days of age, the range of gravity levels used being 2.58-7.38 g. No longevity change was observed for exposures of less than 14 days. The longevity of males increased if they were submitted to hypergravity for durations ranging from 14 to 24 days. This increase in longevity was never observed in females. The positive effect of exposure to hypergravity has been replicated in two laboratories using two wild-type strains and different rearing conditions. A short hypergravity exposure seems to be a mild stress, yielding positive effects on longevity. This is in accordance with two previous studies showing a slight longevity increase after heat shock in the nematode Caenorhabditis elegans and in Drosophila melanogaster.

Animals↗

Effects of hypergravity exposure on the developing central nervous system: possible involvement of thyroid hormone.

The present study examined the effects of hypergravity exposure on the developing brain and specifically explored the possibility that these effects are mediated by altered thyroid status. Thirty-four timed-pregnant Sprague-Dawley rats were exposed to continuous centrifugation at 1.5 G (HG) from gestational Day 11 until one of three key developmental points: postnatal Day (P) 6, P15, or P21 (10 pups/dam: 5 males/5 females). During the 32-day centrifugation, stationary controls (SC, n = 25 dams) were housed in the same room as HG animals. Neonatal body, forebrain, and cerebellum mass and neonatal and maternal thyroid status were assessed at each time point. The body mass of centrifuged neonates was comparatively lower at each time point. The mass of the forebrain and the mass of the cerebellum were maximally reduced in hypergravity-exposed neonates at P6 by 15.9% and 25.6%, respectively. Analysis of neonatal plasma suggested a transient hypothyroid status, as indicated by increased thyroid stimulating hormone (TSH) level (38.6%) at P6, while maternal plasma TSH levels were maximally elevated at P15 (38.9%). Neither neonatal nor maternal plasma TH levels were altered, suggesting a moderate hypothyroid condition. Thus, continuous exposure of the developing rats to hypergravity during the embryonic and neonatal periods has a highly significant effect on the developing forebrain and cerebellum and neonatal thyroid status (P < 0.05, Bonferroni corrected). These data are consistent with the hypothesized role of the thyroid hormone in mediating the effect of hypergravity in the developing central nervous system and begin to define the role of TH in the overall response of the developing organism to altered gravity.

Animals↗

Reduction of G1 phase duration and enhancement of c-myc gene expression in HeLa cells at hypergravity.

We have found that hypergravity stimulates the proliferation of HeLa cells through reduction of the G1 phase duration, concomitant with enhancement of c-myc gene expression. HeLa cells were grown in monolayer in culture flasks that were centrifuged to generate a constant 18, 35 or 70 g at 37 degrees C for up to 4 days. The cell proliferation was enhanced at 18, 35 and 70 g, most notably at 35 g. Cell cycle analyses with [3H]thymidine (TdR)-colcemid treatment showed that the cell generation time in the 35 g culture was reduced by 17% as compared to the control, which was attributed to a 26% reduction of the G1 phase duration. No differences were observed in the duration of the S, G2 and M phases or in the [3H]TdR incorporation per S phase cell between the 35 g culture and the control. The induction of c-myc gene expression was investigated by RNA blot hybridization during a 15-360 min exposure of cells to 18, 35 and 70 g. Elevated levels of c-myc mRNA were observed after a 15-min exposure, and maintained after a 360-min exposure at all hypergravities examined. The highest induction rate of c-myc mRNA was 3.8-fold higher than the control after a 120-min exposure to 35 g. The 35 g condition was the most effective hypergravity for stimulating both cell proliferation and c-myc gene expression. Our study suggests that the appropriate level of hypergravity stimulates HeLa cell proliferation by reducing the G1 phase duration without affecting DNA synthesis rate, mediated through induction of c-myc gene expression.

Cell Cycle↗

Ultrastructure of the parathyroid gland of magnesium-treated golden hamster exposed to a hypergravity environment: a stereological study.

The ultrastructure of the parathyroid glands of magnesium-treated golden hamsters exposed to a 5 gravity environment was studied. In the parathyroid glands of the magnesium-treated animals exposed to a hypergravity environment, the Golgi complexes and cisternae of the granular endoplasmic reticulum were increased as compared to those of the magnesium-treated animals and decreased as compared to those of animals exposed to a hypergravity environment, but were almost the same as those of the control animals. In the control and experimental animals, the chief cells were rich in free ribosomes and mitochondria. In addition, numerous secretory granules were situated close to the plasma membrane in the magnesium-treated animals exposed to a hypergravity environment. These observations suggest that the synthesis of parathyroid hormone may be stimulated in the parathyroid glands of magnesium-treated hamsters exposed to hypergravity environment.

Animals↗

[Effects of hypergravity on migration, proliferation and function of mouse osteoblastic cell line MC3T3-E1].

The purpose of this study was to investigate the hypergravity-induced responses and their mediators of osteoblastic cell line, MC3T3-E1. The synchronized G1 or the S Phase cells were exposed to 5 and 18 x g hypergravity at 37 degrees C. The migration velocity was measured and the morphology was observed. MC3T3-E1 cells were cultured for 1, 2 or 3 days at 37 degrees C, exposing to 5, 10, 20 and 40 x g hypergravity. The proliferation, prostaglandin E2 (PGE2) production rate and alkaline phosphatase (ALPase) activity were measured. The results were as follows: 1) In the G1 phase of the cell cycle, the migration of MC3T3-E1 cells was increased by 18 x g. In the S phase, the morphology altered depending on the g-stress. 2) The proliferation of MC3T3-E1 cells was enhanced at 20 and 40 x g but reduced at 10 x g. The proliferation of HeLa cells and JTC-12 cells was also enhanced at 40 x g. 3) Indomethacin (10(-5)M) reduced the proliferation of MC3T3-E1 cells induced by 40 x g. But indomethacin (10(-5)M) did not reduce the proliferation of HeLa cells. 4) The increase of the released PGE2 from the cells depended on the time (1-8h) and the gravity (1-40 x g). 5) The increase of the ALPase activity of MC3T3-E1 cells also depended on the gravity. These results suggest that the hypergravity enhanced the proliferation of MC3T3-E1 cells via PGE2-mediated mechanism.

Alkaline Phosphatase↗

Electron microscopic study of the parathyroid gland of the calcium-treated hamster subjected to hypergravity environment.

The ultrastructure of the parathyroid glands of calcium-treated golden hamsters subjected to 5 gravity environment was studied. In the calcium-treated animals exposed to hypergravity environment, the Golgi complexes and cisternae of the granular endoplasmic reticulum were significantly decreased compared with those of the animals exposed to hypergravity environment only and appeared to increase compared with those of the calcium-treated animals, but were almost similar to those of the control animals. In addition, many chief cells contained some prosecretory granules in the Golgi areas, some secretory granules situated close to the plasma membrane and many lysosomes. The morphology of the parathyroid glands in the calcium-treated animals exposed to hypergravity environment resembled that of the control animals. These results suggest that the parathyroid glands suppressed by treatment of calcium and stimulated in response to hypergravity environment may indicate the secretory activity of the parathyroid glands of the control animals.

Animals↗

Effects of hypergravity on "whole-blood" cultures of human lymphocytes.

The purpose of this paper is to present a detailed description of the effects of hypergravity on the mitogenic response of human lymphocytes to concanavalin A. The effect on cultures of lymphocytes isolated from peripheral blood are compared with those on whole-blood cultures obtained by diluting fresh blood with culture medium 1:10. Whole-blood cultures of lymphocytes from crew members will be investigated inflight on the Spacelab missions D-1 in 1985 and SLS-1 in 1987. In hypergravity there is an increase in lymphocyte activation of up to 500%. A similar increase can be induced by pre-incubating the cultures in hypergravity prior to exposure to concanavalin A at 1 G. The effect is less evident in cultures of isolated lymphocytes. The influence of autologous plasma and erythrocytes has also been investigated. Plasma and hypergravity have a synergistic and positive effect on lymphocyte activation, i.e. cultures of separated lymphocytes show the highest activation when incubated at 10 G and supplemented with autologous plasma. Conversely, erythrocytes depress lymphocyte activation.

Adaptation, Physiological↗

Hypergravity and estrogen effects on avian anterior pituitary growth hormone and prolactin levels.

Developing female chicks with fractured right radii were maintained for 14 d at either earth gravity (1 G) or a hypergravity state (2 G). The birds at 1 G were divided into groups which received daily injections of 1) saline, 2) 200 micrograms estrone, and 3) 400 micrograms estrone for 14 d. The 2-G birds were divided into three similarly treated groups. All 2-G birds showed significantly lower body weights than did 1-G birds. Anterior pituitary (AP) glands were excise; and analyzed for growth hormone and prolactin content by analytical electrophoresis. The 1-G chicks receiving either dose of daily estrogen showed increased AP growth hormone levels, whereas hypergravity alone did not affect growth hormone content. Chicks exposed to daily estrogen and hypergravity displayed reduced growth hormone levels. AP prolactin levels were slightly increased by the lower daily estrogen dose in 1-G birds, but markedly reduced in birds exposed only to hypergravity. Doubly-treated chicks displayed normal prolactin levels. Reduced growth in 2-G birds might be due, in part, to reduced AP levels of prolactin and/or growth hormone.

Animals↗

The effect of combined hypergravity and micro-grooved surface topography on the behaviour of fibroblasts.

This study evaluated in vitro the differences in morphological behaviour between fibroblast cultured on smooth and micro-grooved substrata (groove depth: 1 mum, width: 1, 2, 5, 10 microm), which undergo artificial hypergravity by centrifugation (10, 24 and 50 g; or 1 g control). The aim of the study was to clarify which of these parameters was more important to determine cell behaviour. Morphological characteristics were investigated using scanning electron microscopy and fluorescence microscopy in order to obtain qualitative information on cell spreading and alignment. Confocal laser scanning microscopy visualised distribution of actin filaments and vinculin anchoring points through immunostaining. Finally, expression of collagen type I, fibronectin, and alpha(1)- and beta(1)-integrin were investigated by PCR. Microscopy and image analysis showed that the fibroblasts aligned along the groove direction on all textured surfaces. On the smooth substrata (control), cells spread out in a random fashion. The alignment of cells cultured on grooved surfaces increased with higher g-forces until a peak value at 25 g. An ANOVA was performed on the data, for all main parameters: topography, gravity force, and time. In this analysis, all parameters proved significant. In addition, most gene levels were reduced by hypergravity. Still, collagen type 1 and fibronectin are seemingly unaffected by time or force. From our data it is concluded that the fibroblasts primarily adjust their shape according to morphological environmental cues like substratum surface whilst a secondary, but significant, role is played by hypergravity forces.

Actin Cytoskeleton↗

Regeneration of guinea pig facial nerve: the effect of hypergravity.

Exposure to moderate hypergravity improves the regenerative capacity of sectioned guinea-pig facial nerve. The improvement in regeneration is tri-directional as follows: a) an average 1.7 fold increase in rate of regeneration in guinea pigs subjected to hypergravity; b) a 25% enhancement of facial muscle activity following the exposure to hypergravity; and c) improvement in the quality of regeneration from an esthetic standpoint. A good correlation was recorded between the histological structure of the severed nerve at the end of the regeneration and the clinical results.

Animals↗

Otoconial alterations after embryonic development in hypergravity.

The relation between prolonged hypergravity and structural adaptation of otoconia was studied in hamsters (n = 56). Three groups of hamsters (n = 27), were conceived and born in a centrifuge: group 1 (n = 10) 1 month under 2.5 G, group 2 (n = 9) 5 months under 2.5 G and 4 months under 1 G, group 3 (n = 8) 1 month under 2.5 G and 8 months under 1 G. Control hamsters (n = 29) were conceived and born under 1 G (1 month old, n = 7; 9 months old, n = 22). Histological study of the otoconial layers (energy dispersive x-ray element analysis and scanning electron microscopy) showed similar calcium content, size, and shape in utricular and saccular otoconia in all groups. Different were the utricular otoconial size classes, large, medium-sized, and small. The area with small otoconia increased in group 1 (p = 0.002). In group 2, the large otoconial area decreased (p = 0.001) and the medium-sized one increased (p < 0.001). In group 3, the large otoconial area decreased (p = 0.003) and the medium-sized one increased (p = 0.007). For age-related effects we found group 1 with an increased area of large otoconia (p = 0.001) and a decreased medium-sized one compared to groups 2 (p < 0.001) and 3 (p = 0.02). Hypergravity during formation of otoconia does not affect calcium content, size, or shape, but changes relative size of the areas with large, medium-sized, or small otoconia and the development of these areas. This resulted in a structural adaptation to hypergravity.

Animals↗

Effect of hypergravity on carboanhydrase reactivity in inner ear ionocytes of developing cichlid fish.

It has been shown earlier that hypergravity slows down inner ear otolith growth in developing fish. Otolith growth in terms of mineralization mainly depends on the enzyme carboanhydrase (CA), which is responsible for the provision of the pH-value necessary for calcium carbonate deposition. Larval siblings of cichlid fish (Oreochromis mossambicus) were subjected to hypergravity (3 g, hg; 6 h) during development and separated into normally and kinetotically swimming individuals following the transfer to 1 g (i.e., stopping the centrifuge; kinetotically behaving fish performed spinning movements). Subsequently, CA was histochemically demonstrated in inner ear ionocytes (cells involved in the endolymphatic ion exchange) and enzyme reactivity was determined densitometrically. It was found that both the total macular CA-reactivity as well as the difference in reactivities between the left and the right maculae (asymmetry) were significantly lower (1) in experimental animals as compared to the 1 g controls and (2) in normally swimming hg-animals as compared to the kinetotically behaving hg-fish. The results are in complete agreement with earlier studies, according to which hypergravity induces a decrease of otolith growth and the otolithic calcium incorporation (visualized using the calcium-tracer alizarin complexone) of kinetotically swimming hg-fish was higher as compared to normally behaving hyper-g animals. The present study thus strongly supports the concept that a regulatory mechanism, which adjusts otolith size and asymmetry as well as otolithic calcium carbonate incorporation towards the gravity vector, acts via activation/deactivation of macular CA.

Animals↗

Hypergravity induces expression of cyclooxygenase-2 in the heart vessels.

Cyclooxygenase-2 (COX-2), a rate-limiting enzyme for prostaglandin biosynthesis, is induced by various stimuli including mechanical stress and plays important roles in pathophysiological conditions. For example, gravitational stress has been shown to induce expression of COX-2 in bone tissues, which is essential for bone homeostasis. To investigate whether COX-2 is induced by gravitational loading in other tissues than bone, we exposed mice to hypergravity at 2G and 3G for 4 h. We demonstrate here that COX-2 is induced in the mouse heart vessels by hypergravity. Moreover, hypoxia-inducible factor (HIF)-1alpha and its downstream genes such as inducible nitric oxide synthase, vascular endothelial growth factor, and heme oxygenase-1 were induced in the heart simultaneously, while none of these genes were induced in the COX-2(-/-) mouse heart. Therefore, COX-2 induced in the heart helps protect the heart function against hypoxia under hypergravity condition through HIF-1alpha induction.

Animals↗