Research Containing: Hindlimb Suspension

Impaired osteoblastogenesis potential of progenitor cells in skeletal unloading is associated with alterations in angiogenic and energy metabolism profile

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Skeletal unloading provokes bone loss. These bone alterations have been shown to be associated with impairment of osteoblastic activity. In the present study, we evaluated the effect of skeletal unloading on bone marrow progenitor cells, for exploration of the underlying mechanism. Wistar rats were randomized to be either hindlimb unloaded for 9 days or to act as controls. Micro-CT was used to detect tibial trabecular architecture changes in response to skeletal unloading. Microgravity conditions for 9 days resulted in a decreased number and an increased spacing of the bone trabeculae in the proximal tibia. The proliferative capacity of the femoral bone marrow samples was assessed (fibroblast-colony-forming assay). By using qPCR, the expression of selected markers of vascularization (Vegfa; Hif1a; Angpt1), energy metabolism (Prkaa2; Mtor), bone formation (Runx2; Alp; Bglap; Bmp2; Bmp4; Bmp7) and bone resorption (Acp5; Tnfsf11; Tnfrsf11b) in these bone marrow suspensions was measured. We demonstrated a striking decrease in the number of fibroblastic progenitors in response to hindlimb unloading. This deficit in proliferation was shown to be accompanied by altered hindlimb perfusion and cellular energy homeostasis. Ex vivo culture assays of the bone marrow-derived progenitor cells screened for osteogenic (Runx2; Alp; Bglap) and adipogenic (Pparg; Fabp4) differentiation alterations in response to microgravity. Induced progenitor cells from unloaded rats showed a delay in osteogenic differentiation and impaired adipogenic differentiation compared to control. The data of this multi-level approach demonstrate that skeletal unloading significantly affects the bone tissue and its metabolism at the progenitor stage. The molecular expressions of the bone marrow population support a role of cellular metabolic stresses in skeletal alterations induced by inactivity.

Related URLs:
<Go to ISI>://WOS:000306372100004

Long-term (6-wk) hindlimb suspension inhibits spermatogenesis in adult male rats

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The International Space Station will allow extended habitation in space and long-term exposure to microgravity (microG). A concern is the impact of long-term microG exposure on the ability of species to reproduce. The model often used to simulate microG is rat hindlimb suspension (HLS), where the hindlimbs are elevated above the cage floor with a tail harness. Experiments described here are the first to examine the effect of long-term HLS on testicular function in adult male rats. Free-roaming (controls), animals with only the tail harnessed but hindlimbs in contact with the cage floor (TO), and HLS animals were tested for 6 wk. Cryptorchidism was prevented in TO and HLS animals by partial constriction of the inguinal canal with sutures. All parameters were compared at the end of the 6-wk experiment. Testicular weights and spermatogenesis were significantly reduced by HLS, such that no spermatogenic cells beyond round spermatids were present and epididymides were devoid of mature sperm. In many tubules, loss of all germ cells, except a few spermatogonia, resulting in histopathology similar to the Sertoli cell, was observed. Spermatogenesis appeared unaffected in control and TO animals. Sertoli and Leydig cell appearance, testosterone, luteinizing hormone, and follicle-stimulating hormone levels, and epididymal and seminal vesicle weight were unchanged by HLS. Cortisone was not elevated by HLS; thus stress may not be a factor. These results demonstrate that spermatogenesis is severely inhibited by long-term HLS, whereas testicular androgen production is not. These results have significant implications regarding serious effects of long-term exposure to microG on the reproductive capability of scrotal mammals, including humans.

Biomedical analysis of rat body hair after hindlimb suspension for 14 days

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The levels of 26 minerals in rat body hair were analyzed in control and hindlimb-suspended Wistar Hannover rats (n=5 each). We quantified the levels of 22 minerals in this experiment. However, we were unable to measure the levels of 4 minerals (Be, V, Cd, and Hg) quantitatively because they were below the limit of detection. Of the 22 quantified, the levels of 19 minerals were not significantly different between control and hindlimb-suspended groups. The levels of 3 minerals (Pb, Cr, and Al) tended to be higher in the hindlimb-suspended group than in the control group; however, this difference was not significant. The concentrations of 3 other minerals (I, K, and Mg) were significantly different between the 2 groups. The iodine (I) level was 58.2% higher in the hindlimb-suspended group than in the control group (P<0.05). Potassium (K) and magnesium (Mg) levels were 55.2% and 20.4% lower, respectively, in the experimental group (0.05 in both cases). These results indicate that a physiological change in mineral metabolism resulting from physical or mental stress, such as hindlimb suspension, is reflected in body hair. The Japan Aerospace Exploration Agency (JAXA) has initiated a human research study to investigate the effects of long-term space flight on gene expression and mineral metabolism by analyzing hair samples of astronauts who stayed in the International Space Station (ISS) for 6 months. We believe that hindlimb suspension for 14 days can simulate the effects of an extremely severe environment, such as space flight, because the hindlimb suspension model elicits a rapid physiological change in skeletal muscle, bone, and fluid shift even in the short term. These results also suggest that we can detect various effects on the body by analyzing the human scalp hair shaft.

Simulation models of weightlessness in mammalian's developmental program

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Effects of gravitational loading levels on protein expression related to metabolic and/or morphologic properties of mouse neck muscles

by cfynanon 9 June 2015in Biology & Biotechnology No comment

The effects of 3 months of spaceflight (SF), hindlimb suspension, or exposure to 2G on the characteristics of neck muscle in mice were studied. Three 8‐week‐old male C57BL/10J wild‐type mice were exposed to microgravity on the International Space Station in mouse drawer system (MDS) project, although only one mouse returned to the Earth alive. Housing of mice in a small MDS cage (11.6 × 9.8‐cm and 8.4‐cm height) and/or in a regular vivarium cage was also performed as the ground controls. Furthermore, ground‐based hindlimb suspension and 2G exposure by using animal centrifuge (n = 5 each group) were performed. SF‐related shift of fiber phenotype from type I to II and atrophy of type I fibers were noted. Shift of fiber phenotype was related to downregulation of mitochondrial proteins and upregulation of glycolytic proteins, suggesting a shift from oxidative to glycolytic metabolism. The responses of proteins related to calcium handling, myofibrillar structure, and heat stress were also closely related to the shift of muscular properties toward fast‐twitch type. Surprisingly, responses of proteins to 2G exposure and hindlimb suspension were similar to SF, although the shift of fiber types and atrophy were not statistically significant. These phenomena may be related to the behavior of mice that the relaxed posture without lifting their head up was maintained after about 2 weeks. It was suggested that inhibition of normal muscular activities associated with gravitational unloading causes significant changes in the protein expression related to metabolic and/or morphological properties in mouse neck muscle. mouse neck musclegravitational unloadingprotein expression.

The current state of bone loss research: Data from spaceflight and microgravity simulators

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Bone loss is a well documented phenomenon occurring in humans both in short- and in long-term spaceflights. This phenomenon can be also reproduced on the ground in human and animals and also modeled in cell-based analogs. Since space flights are infrequent and expensive to study the biomedical effects of microgravity on the human body, much of the known pathology of bone loss comes from experimental studies. The most commonly used in vitro simulators of microgravity are clinostats while in vivo simulators include the bed rest studies in humans and hindlimb unloading experiments in animals. Despite the numerous reports that have documented bone loss in wide ranges in multiple crew members, the pathology remains a key concern and development of effective countermeasures is still a major task. Thus far, the offered modalities have not shown much success in preventing or alleviating bone loss in astronauts and cosmonauts. The objective of this review is to capture the most recent research on bone loss from spaceflights, bed rest and hindlimb unloading, and in vitro studies utilizing cellular models in clinostats. Additionally, this review offers projections on where the research has to focus to ensure the most rapid development of effective countermeasures.

Related URLs:
http://dx.doi.org/10.1002/jcb.24454

Spaceflight regulates ryanodine receptor subtype 1 in portal vein myocytes in the opposite way of hypertension

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Gravity has a structural role for living systems. Tissue development, architecture, and organization are modified when the gravity vector is changed. In particular, microgravity induces a redistribution of blood volume and thus pressure in the astronaut body, abolishing an upright blood pressure gradient, inducing orthostatic hypotension. The present study was designed to investigate whether isolated vascular smooth muscle cells are directly sensitive to altered gravitational forces and, second, whether sustained blood pressure changes act on the same molecular target. Exposure to microgravity during 8 days in the International Space Station induced the decrease of ryanodine receptor subtype 1 expression in primary cultured myocytes from rat hepatic portal vein. Identical results were found in portal vein from mice exposed to microgravity during an 8-day shuttle spaceflight. To evaluate the functional consequences of this physiological adaptation, we have compared evoked calcium signals obtained in myocytes from hindlimb unloaded rats, in which the shift of blood pressure mimics the one produced by the microgravity, with those obtained in myocytes from rats injected with antisense oligonucleotide directed against ryanodine receptor subtype 1. In both conditions, calcium signals implicating calcium-induced calcium release were significantly decreased. In contrast, in spontaneous hypertensive rat, an increase in ryanodine receptor subtype 1 expression was observed as well as the calcium-induced calcium release mechanism. Taken together, our results shown that myocytes were directly sensitive to gravity level and that they adapt their calcium signaling pathways to pressure by the regulation of the ryanodine receptor subtype 1 expression.

Effects of pleiotrophin overexpression on mouse skeletal muscles in normal loading and in actual and simulated microgravity

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Pleiotrophin (PTN) is a widespread cytokine involved in bone formation, neurite outgrowth, and angiogenesis. In skeletal muscle, PTN is upregulated during myogenesis, post-synaptic induction, and regeneration after crushing, but little is known regarding its effects on muscle function. Here, we describe the effects of PTN on the slow-twitch soleus and fast-twitch extensor digitorum longus (EDL) muscles in mice over-expressing PTN under the control of a bone promoter. The mice were maintained in normal loading or disuse condition, induced by hindlimb unloading (HU) for 14 days. Effects of exposition to near-zero gravity during a 3-months spaceflight (SF) into the Mice Drawer System are also reported. In normal loading, PTN overexpression had no effect on muscle fiber cross-sectional area, but shifted soleus muscle toward a slower phenotype, as shown by an increased number of oxidative type 1 fibers, and increased gene expression of cytochrome c oxidase subunit IV and citrate synthase. The cytokine increased soleus and EDL capillary-to-fiber ratio. PTN overexpression did not prevent soleus muscle atrophy, slow-to-fast transition, and capillary regression induced by SF and HU. Nevertheless, PTN exerted various effects on sarcolemma ion channel expression/function and resting cytosolic Ca(2+) concentration in soleus and EDL muscles, in normal loading and after HU. In conclusion, the results show very similar effects of HU and SF on mouse soleus muscle, including activation of specific gene programs. The EDL muscle is able to counterbalance this latter, probably by activating compensatory mechanisms. The numerous effects of PTN on muscle gene expression and functional parameters demonstrate the sensitivity of muscle fibers to the cytokine. Although little benefit was found in HU muscle disuse, PTN may emerge useful in various muscle diseases, because it exerts synergetic actions on muscle fibers and vessels, which could enforce oxidative metabolism and ameliorate muscle performance.

Effect of simulated weightlessness on osteoprogenitor cell number and proliferation in young and adult rats

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Experiments with rats flown in space or hind limb unloaded (HU) indicate that bone loss in both conditions is associated with a decrease in bone volume and osteoblast surface in cancellous and cortical bone. We hypothesize that the decrease in osteoblastic bone formation and osteoblast surface is related to a decrease in the number of osteoprogenitors and/or decreased proliferation of their progeny. We tested this hypothesis by evaluating the effect of 14 days of HU on the number of osteoprogenitors (osteoblast colony forming units; CFU-O), fibroblastic colony forming units (CFU-F), and alkaline phosphatase-positive CFU (CFU-AP) in cell populations derived from the proximal femur (unloaded) and the proximal humerus (normally loaded) in 6-week-old and 6-month-old rats. To confirm the effect of unloading on bone volume and structure, static histomorphometric parameters were measured in the proximal tibial metaphysis. Effects of HU on proliferation of osteoprogenitors were evaluated by measuring the size of CFU-O. HU did not affect the total number of progenitors (CFU-F) in young or adult rats in any of the cell populations. In femoral populations of young rats, HU decreased CFU-O by 71.0% and mean colony size was reduced by 20%. HU decreased CFU-AP by 31.3%. As expected, no changes in CFU-O or CFU-AP were seen in cell populations from the humerus. In femoral cell populations of adult rats, HU decreased CFU-O and CFU-AP by 16.6% and 36.6%, respectively. Again, no effects were seen in cell populations from the humerus. In 6-week-old rats, there was a greater decrease in bone volume, osteoblast number, and osteoblast surface in the proximal tibial metaphysis than that observed in adult rats. Both trabecular thickness and trabecular number were decreased in young rats but remained unaffected in adults. Neither osteoclast number nor surface was affected by unloading. Our results show that the HU-induced decrease in the number of osteoprogenitors observed in vitro parallels the effects of HU on bone volume and osteoblast number in young and old rats in vivo, suggesting that the two may be interdependent. HU also reduced CFU-O colony size in femoral populations indicating a diminished proliferative capacity of osteoblastic colonies.

Effects of spaceflight on murine skeletal muscle gene expression

by cfynanon 9 June 2015in Biology & Biotechnology No comment

Spaceflight results in a number of adaptations to skeletal muscle, including atrophy and shifts toward faster muscle fiber types. To identify changes in gene expression that may underlie these adaptations, we used both microarray expression analysis and real-time polymerase chain reaction to quantify shifts in mRNA levels in the gastrocnemius from mice flown on the 11-day, 19-h STS-108 shuttle flight and from normal gravity controls. Spaceflight data also were compared with the ground-based unloading model of hindlimb suspension, with one group of pure suspension and one of suspension followed by 3.5 h of reloading to mimic the time between landing and euthanization of the spaceflight mice. Analysis of microarray data revealed that 272 mRNAs were significantly altered by spaceflight, the majority of which displayed similar responses to hindlimb suspension, whereas reloading tended to counteract these responses. Several mRNAs altered by spaceflight were associated with muscle growth, including the phosphatidylinositol 3-kinase regulatory subunit p85alpha, insulin response substrate-1, the forkhead box O1 transcription factor, and MAFbx/atrogin1. Moreover, myostatin mRNA expression tended to increase, whereas mRNA levels of the myostatin inhibitor FSTL3 tended to decrease, in response to spaceflight. In addition, mRNA levels of the slow oxidative fiber-associated transcriptional coactivator peroxisome proliferator-associated receptor (PPAR)-gamma coactivator-1alpha and the transcription factor PPAR-alpha were significantly decreased in spaceflight gastrocnemius. Finally, spaceflight resulted in a significant decrease in levels of the microRNA miR-206. Together these data demonstrate that spaceflight induces significant changes in mRNA expression of genes associated with muscle growth and fiber type.

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Research Containing: Hindlimb Suspension

Impaired osteoblastogenesis potential of progenitor cells in skeletal unloading is associated with alterations in angiogenic and energy metabolism profile

Long-term (6-wk) hindlimb suspension inhibits spermatogenesis in adult male rats

Biomedical analysis of rat body hair after hindlimb suspension for 14 days

Simulation models of weightlessness in mammalian's developmental program

Effects of gravitational loading levels on protein expression related to metabolic and/or morphologic properties of mouse neck muscles

The current state of bone loss research: Data from spaceflight and microgravity simulators

Spaceflight regulates ryanodine receptor subtype 1 in portal vein myocytes in the opposite way of hypertension

Effects of pleiotrophin overexpression on mouse skeletal muscles in normal loading and in actual and simulated microgravity

Effect of simulated weightlessness on osteoprogenitor cell number and proliferation in young and adult rats

Effects of spaceflight on murine skeletal muscle gene expression