Mice Drawer System (MDS) - 03.24.16
Mice Drawer System (MDS) is an Italian Space Agency experiment that will use a validated mouse model to investigate the genetic mechanisms underlying bone mass loss and other microgravity effect on different tissues such as muscles, glands, brain. Research conducted with the MDS is an analog to the human research program, which has the objective to extend the human presence safely beyond low Earth orbit. Science Results for Everyone
The Mice Drawer System (MDS) used three wild-type (Wt) and three PTN mice (which have had a foreign bone-metabolism gene inserted) to investigate genetic mechanisms of bone loss and effects on other tissues in microgravity. One PTN and two Wt mice died due to health- or payload-related reasons, but tissues were used in 20 different investigations of microgravity-induced modifications, primarily focusing on bone loss. These revealed loss in the weight-bearing bones of both strains; muscle atrophy; changes in atrophy-related genes; structural changes in thyroid and testes; changes in regulation of immune response, metabolic, inflammatory and catabolic genes; and changes in blood cell parameters. Experiment Details
Ranieri Cancedda, M.D., University of Genoa (Unige) and IRCCS AOU San Martino – IST National Cancer Research Institute, Genoa, Italy
Francesco Saverio Ambesi-Impiombato, University of Udine, Udine, Italy
Angela Maria Rizzo, Ph.D., University of Milan, Milan, Italy
Antonio Musaro, University of Rome La Sapienza, Rome, Italy
Betty Nusgens, Prof., University of Liege, Sart-Tilman, Belgium
Charles A. Fuller, University of California, Davis, Davis, CA, United States
Yoshinobu Ohira, Doshisha University, Kyoto, Japan
Daniela Santucci, Istituto Superiore di Sanita’, Roma, Italy
Diana Conte Camerino, University of Bari, Bari, Italy
Dieter Blottner, Ph.D., Charite Campus Benjamin Franklin, Berlin, Germany
Akihiko Ishihara, Kyoto University, Kyoto, Japan
Felice Strollo, Instituto Nazionale Riposo e Cura Anziani (INRCA), Rome, Italy
Giuseppe Miserocchi, University of Milano, Milan, Italy
Jean-Luc Morel, Physiologie Cellulaire et Pharmacologie Moleculaire, Bordeaux, France
Michael J. Pecaut, Ph.D., Loma Linda University, CA, United States
Paolo Di Nardo, University of Rome Tor Vergata, Rome, Italy
Sara Tavella, Università degli Studi di Genova, Genova, Italy
Alessandra Ruggiu, University of Genoa (Unige) and National Cancer Research Institute, Genova, Italy
Proto Pippia, Universiy of Sassari, Sassari, Italy
Stefano Schiaffino, University of Padua, Padua, Italy
Ted A. Bateman, Ph.D., University of North Carolina, Chapel Hill, NC, United States
Yi Liu, Ph.D., University of Genoa (Unige) and National Cancer Research Institute, Genoa, Italy
Yufang Shi, Ph.D., University of Medicine and Dentistry of New Jersey, Newark, NJ, United States
Thales Alenia Space, Milan, Italy
Sponsoring Space Agency
National Aeronautics and Space Administration (NASA)
Italian Space Agency (ASI)
ISS Expedition Duration 1
April 2009 - March 2010; March 2014 - September 2014
Previous ISS Missions
MDS is a new investigation for ISS research.
- The Mice Drawer System (MDS) is an Italian Space Agency facility developed by Thales Alenia Space-Italia in order to support up to six mice onboard the ISS during long-duration exploration missions lasting 100 days with a potential eighty day extension.
- While in space, crewmembers do not experience mechanical stimuli inducing physiological changes in bone architecture and functionality resulting in osteoporotic-like effects which are similar to age-related osteoporosis.
- Pleiotrophin transgenic mice (PTN-Tg) were selected because this transgene (a foreign gene that has been inserted into its genome to exhibit a particular trait) plays an important role in bone metabolism. MDS tests the hypothesis that mice with an overexpression of the PTN gene will be protected from the osteoporosis-like effects due to microgravity.
- MDS examines a wide range of physiological systems, such as muscle, bone, and blood, along with different organs and glands by compiling an integrative view of the mammal’s physiological responses to microgravity. In effort to understand the effects of microgravity on specific tissue samples, a Tissue Sharing Program consisting of 20 research groups representing 6 countries was assembled.
Almost every function of the human body is affected by exposure to microgravity and typically involves complex changes in human tissues and organs. However, in most cases, the adaptions induced by microgravity are only temporary and typically return to normal after returning to Earth.
MDS was designed to employ mice as the animal model utilizing transgenic mice with an overexpression of the pleiotropin (PTN) gene that is under the control of the same specific human bone promoter of the osteocalcin gene. This gene was selected because of the positive effect it has on bone metabolism. PTN, also known as osteoblast stimulating factor-1 (OSF-1), is an extracellular matrix-associated growth/differentiation factor. PTN defines a new family of secreted heparin-binding growth factors structurally unrelated to other growth factor families. This growth factor shows different functions, ranging from stimulating neurogenesis (process of nerve formation), cell proliferation, and chemotaxis (when cells orient their movements toward certain environmental chemicals) to tumor angiogenesis (creation of new blood vessels to a tumor).
MDS was carried to the ISS on August 28, 2009, and returned on November 27th, 2009, setting the longest permanence in space for rodents of 91 days. Three wild type (Wt) and three PTN-transgenic male mice were housed in the MDS during this experiment. A ground replica mimicking the flight experiment was performed at the University of Genova to evaluate the effects of microgravity and compare these effects to different Earth controls.
Unfortunately during the mission, one PTN-Tg and two Wt mice died. The necropsy revealed that mouse Wt3 had a major spinal cord lesion that possibly occurred during the shuttle lift off. An analysis performed on feces present in the cage of the PTN-Tg3 mouse suggested that the animal could have developed a liver pathology. The post-landing check performed on the mice cages revealed that Wt1 mouse died in consequence of a failure of the food cassette system. The remaining mice showed a normal behavior throughout the experiment and appeared in excellent health conditions after landing. During the mission, the mice health conditions and their water and food consumption were checked daily. Upon landing, mice were sacrificed, blood parameters were measured and tissues were dissected for subsequent analysis.
The collected samples deriving from almost every tissue of the mice body were analysed by using different techniques, such as:
- Histomorphometry measurements
- Imaging with x-ray microtomography
- Molecular studies
- Biochemical analysis
- Gene expression analysis
This research is also expected to contribute data to the current body of research on microgravity effects on the skeletal, cardiovascular, and immune systems, liver and kidney function as well as other physiological systems through a tissue sharing program. Every effort will be made to harvest as many different samples and types of tissue from the mice as possible for other mission specific biomedical research. Positive results from this research may advance our understanding of mechanistic changes that occur in various physiological systems after exposure to microgravity and support overall efforts to reduce health risks to crewmembers. The experiment’s resulting from the MDS tissue sharing program are as follows:
A transgenic approach to space osteoporosis
Ranieri Cancedda, University of Genova, Liguria, Italy
Tissue: Bone, Blood
Investigate the effects of microgravity on transgenic (inbred mice) as a tool to study genetic mechanisms underlying bone mass pathophysiology (study of the disturbance of normal mechanical, physical, and biochemical functions caused by a disease). The project is based on the rationale that mice with an increased bone density are likely to be better protected from osteoporosis, when their phenotype is a direct effect of gene(s) involved in skeletogenesis (process to produce the skeleton).
Akt-FoxO signaling and protein degradation pathways during muscle atrophy induced by space flight
Stefano Schiaffino, University of Padova, Padua, Italy
Tissue: Anterior tibialis, Gastrocnemius and other muscles
Muscle wasting occurs in many conditions such as disuse, fasting, cancer, diabetes and renal failure. Space flight is known to cause muscle atrophy. However, the activity of signaling pathways during space flights has never been explored. We will investigate the role of Akt-FoxO signaling pathway as the main cause of muscle loss during space flight.
Study of the IGF-1 system in mice subjected to space flight
Antonio Musaro, University of Rome La Sapienza, Rome Italy
Tissue: Soleus, Extensor digitorum longus, Anterior tibialis
Investigate whether the IGF-1 signaling pathway is affected in the skeletal muscle of mice subjected to space flight. In particular, soon after the mice return to the Earth, the different muscle will be isolated and immediately frozen in liquid nitrogen and then stored at -70 degrees C until analysis. The gene and protein expression levels of muscle IGF-1, IGF-1R will be assessed in all muscle biopsies and correlated with histological analysis. This analysis will allow correlation to the expression of mIGF-1 as a function of weight loss. This study will also evaluate whether space flight affects specific signal transduction pathways that mediate the IGF-1 actions, such as PI3-Kinase/Akt, MAPK, calcineurin. This study will also determine the potential signal transduction pathways associated with muscle atrophy such as the ubiquitin ligase, atrogin-1/MAFbx, and Foxo. The results obtained will provide useful data to assess the status of IGF-1 pathway in the muscle of mice subjected to space flight; design specific therapeutic tools to counteract muscle atrophy associated to space flight; reduce muscle atrophy in future space explorers.
Effects of microgravity on ion channel function and expression in mouse skeletal muscle: role in disuse-induced muscle atrophy and phenotype transition, and potential targets for therapeutic intervention
Diana Conte Camerino, University of Bari, Bari, Italy
Tissue: Soleus, Extensor digitorum longus, Anterior tibialis, Urine filter
Using the HU model of simulated microgravity, it appeared that ion channel expression/function is changed by muscle disuse in relation to muscle phenotype transition and/or muscle atrophy, thereby contributing to muscle adaptation/alteration and constituting possible therapeutic targets. The opportunity offered by the MDS facility to study ion channel changes occurring during actual microgravity is very important for two main reasons. First, it is critical to verify that the effects observed with the HU model occur during space flight, in order to further validate the HU mouse as a valuable model of simulated microgravity. Second, the MDS facility will offer a huge opportunity to test drugs acting on sarcolemma ion channels as potential countermeasures against the muscle alteration induced by microgravity, and more generally by Earth conditions such as muscle disuse/immobilization, inherited and acquired muscle dystrophies, or ageing.
Effects of space flight on erythrocytes and oxidative stress of rodents
Angela Maria Rizzo, University of Milano, Milan, Italy
Erythrocyte and hemoglobin loss has been continuously observed during space missions; these observations have been summarized as “space anemia”. Many studies have demonstrated that erythrocytes exposed to microgravity whether in vivo or in vitro, have a modified rheology and undergo greater hemolysis. We can suppose that microgravity together with space radiation causes variations of cellular shape, plasma membrane composition, and peroxidative stress, that can be responsible of space anemia. The availability of rodents blood and tissues through a biospecimen sharing plan is a good opportunity to investigate the in vivo effects of space environment on erythrocytes, androgens levels and oxidative stress. The aim of our project is to analyze red blood cell membrane composition and to determine the oxidative stress that these animals and their erythrocytes have undergone.
Endocrine determinants of rodent musculo-skeletal ageing phenomena in space
Felice Strollo, Instituto Nazionale Riposo e Cura Anziani (INRCA), Rome, Italy
Tissue: Gonads, Hypophysis, Stomach, Urine filter
Microgravity causes osteoporosis by an ageing-like mechanism in males and females. In fact, impaired osteogenesis might contribute to it more than enhanced osteolysis, as it happens with ageing. A parallel decrease in muscle mass occurs in space. The aim of this study is to try and go deeper into the overall underlying endocrine-related mechanisms of these phenomena by correlating brain and gonadal function changes with some drivers of metabolic adaptation potentially involved in such changes, including visceral adiposity and circadian rhythm.
Mouse behavioural observation of mouse under microgravity condition ethogram definition and neurobiological correlates
Daniela Santucci, Istituto Superiore di Sanita, Rome, Italy
Tissue: Brain, Adrenal Glands, Tongue
The aim of the proposed research is the study of role of gravitational environment in mammalian neurobehavioral response. Increasing evidence from both ground-based and space research indicate that nervous system is potentially affected by exposure to hyper/microgravity. We intend to evaluate changes in neurochemical, neuromorphological, hormonal, and behavioral parameters during and following exposure to space flight in mice. In particular, video recorded data obtained by the OSS available on the MDS facility during the experiment will be analyzed in order to finely define the behavioral repertoire (ethogram) of the mice exposed to space environment. Moreover, neurobiological correlates known to be involved in the responses to stress, such as blood or tissue levels of NGF, BDNF, interleukin, and relative receptors, will be evaluated in brain, adrenal glands, tongue and limbs.
Role of stem cells in cardiac muscle deconditioning in microgravity
Paolo Di Nardo, University of Rome Tor Vergata, Rome, Italy
Tissue: Myocardium, Diaphragm
It has been demonstrated that adult cardiomyocytes can be progressively substituted within the myocardium by newly formed contractile cells originated from stem cells resident in the heart and/or recruited from the blood stream, either in the form of the endogenous cells or following systemic infusion from a donor. This implies that, in microgravity, muscle atrophy could be ascribed, among other factors, to an impaired stem cell/cardiomyocyte turnover. Therefore, the present experiment is aimed at evaluating whether microgravity could modify stem cell activation within the mammalian myocardium. To this end, in a first step, the number and molecular and biochemical features of stem cells embedded in the myocardium of mice exposed to microgravity will be assessed.
Lungs genomics in mice chronically exposed to microgravity
Giuseppe Miserocchi, University of Milano, Milan, Italy
The Mice Drawer System (MDS) offers the opportunity to expose mice to long-duration microgravity. This study will evaluate the functional conditions of the lungs after chronic microgravity exposure. This research will be performed based on previous studies on extravascular matrix in experimental interstitial lung edema. This study will develop high resolution morphometry of the lungs; perform gene expression analysis of various pro-inflammatory cytokines and possibly whole transcriptome analysis using microarray; estimate signalling in endothelial and epithelial cells; evaluate the modifications induced on the interstitial macromolecular structure of the extracellular matrix. The integration of data from these four study lines should allow a thorough evaluation of the impact of chronic microgravity exposure on the respiratory function.
Effect of real microgravity on the expression of proteins involved in the rat intestinal transit
Proto Pippia, Universiy of Sassari, Sassari, Italy
Tissue: Stomach and Intestine
The modification or the loss of the gravitational force vector strongly affects many physiological functions as a result of biological process modifications. Many space missions have shown that prolonged exposure of humans to extended weightlessness may seriously affect their health. The modifications of physiological parameters seem to be a direct consequence of changes in cellular activities, as well as protein expression in many processes involving the regulation of cell growth, metabolism, signal transduction and transcription, apoptosis and tumor suppression. Numerous studies have revealed changes in gastrointestinal secretion, motility, evacuation and absorption in humans and animals after space missions: among biological alterations, low gravity generates modification of gastrointestinal function as enzyme activity, intestinal mucosa integrity, colonic microflora, and liver activity. The aim of the study is to determine whether real microgravity influence the expression of enzymes involved in the intestinal transit and gastrointestinal homeostasis as the inducibile isoform of nitric oxide synthase (iNOS), ciclooxygenase (COX-1 and COX-2), ICAM-1 and heat shock proteins 70 (HSP 70) and 90 (HSP 90).
Effects of Microgravity on heart mass and Extracellular Matrix
Michael D. Delp, Ph.D., University of Florida, Gainesville, FL
It is hypothesized that microgravity will induce cardiac atrophy in the flight animal. Total heart mass, as well as right ventricular, left ventricular and septal mass will be measured and compared to that of ground-based vivarium control and animal enclosure module (AEM) control mice. To determine whether microgravity alters cardiac extracellular matrix protein concentration and composition. It is hypothesized that microgravity will decrease total collagen content in the right ventricle, left ventricle and septum; alter the collagen composition of the heart toward the isoform that would favor increased cardiac compliance, i.e., less type I collagen and more type III collagen, and/or decrease the cross-liking among collagen fibers.
The effect of space flight on the immune system and stress response in osteoblast stimulating factor-1 transgenic mice
Yufang Shi, Ph.D., University of Medicine and Dentistry of New Jersey, Newark, NJ
Tissue: Thymus, spleen and serum
Using the mouse hindlimb suspension model, we have found that mice subjected to this condition have significant reductions in lymphocyte numbers, which occurs through apoptosis. These mice also exhibited significant increases in serum levels of haptoglobin and corticosteroid. Increased serum haptoglobin has also been observed in crewmembers during space flight, and it is currently being evaluated as a marker for stress levels. Recently, we found that mice deficient in osteopontin (OPN) are resistant to hindlimb unloading-induced increases in corticosteroids. Therefore, we would like to verify these observations in mice flown in space. Since osteoblast stimulating factor-1 (OSF1) has been shown to be an important cell survival factor, we believe that overexpression of OSF1 may affect the survival of lymphocytes. In addition, with the increased bone formation in OSF1 transgenic mice, it is possible that there will be less OPN released from bone under conditions of microgravity during space flight, thus reducing corticosteroid production. Therefore, we will examine the extent of depletion of lymphocytes, and the changes in haptoglobin and corticosteroid levels in wild type and OSF1-transgenic mice immediately after space flight.
Cellular/molecular causes of skeletal deterioration
Ted A. Bateman, Ph.D., Clemson University, Clemson, SC
Tissue: Long bones, Spine
We would analyze skeletal tissue by assays similar to those used for examining bones from mice flown on CBTM-01 and CBTM-02 payloads. This includes 1) microcomputed tomography (microCT) of both trabecular and cortical bone from multiple sites. As a non-destructive test microCT can proceed and even direct future analysis such as histology, quantitative histomorphometry, mechanical testing, or material property studies. 2) mechanical testing of the femur diaphysis in three- (or four-) point bending. 3) compositional analysis of mineral and organic constituents (this can be performed on non-traditional skeletal tissue like ribs). 4) histology and histomorphometry of decalcified (paraffin and glycol methacrylate techniques) or undecalcified section. 5) Immunohistochemistry of decalcified embedded sections for apoptosis, macrophage activation, inflammation, and osteoclast activation proteins with an emphasis on the RANK/RANKL/OPG pathway. 6) ELISA/CBA analysis of serum for bone formation, bone resorption and inflammatory markers.
Effects of long-duration space flight on the circadian and metabolic systems of mice
Charles A. Fuller, University of California - Davis, Davis, CA
Tissue: Eyes, Liver Epididymal fat pad, Brain, Plasma
Circadian rhythms and body mass regulation/energy metabolism are critical homeostatic systems for the health and well being of organisms. Further, these systems have been demonstrated to be influenced by altered gravity, including space flight. This program is requesting tissues and data to help understand the effects of long-duration space flight on the regulation of these systems. The hypotheses we propose to test are:
- Mice exposed to 100 days of microgravity will, compared to ground controls, demonstrate altered retinal and hypothalamic morphology, circadian clock gene activity, and reduced sensitivity to light exposure, the primary environmental stimulus for the circadian system.
- Mice exposed to 100 days of microgravity will, compared to ground controls, demonstrate increased body and fat mass, decreased food intake, altered levels of hypothalamic neuropeptides related to energy balance, and liver function will be shifted to support fat storage.
Effects of space flight on genes/proteins ex in brain/muscle
Yoshinobu Ohira, Osaka University
Tissue: Brain, Muscle, Plasma, Testis
This experiment will be performed to investigate the adaptation of brain, adductor longus muscle, testis, and blood plasma of rats to gravitational unloading by actual space flight and/or simulation model, hindlimb suspension. The major parameters to be analyzed are expression of genes and proteins in various regions of brain and adductor longus muscle and hormones in blood plasma. As for the adductor longus muscle, histochemical analyses, including the size and number of myonuclei and satellite cells, DNA content and number of nucleoli in each myonucleus, cross-sectional area, and length of fibers and specific gene expression, such as heat shock protein, will be also analyzed in longitudinal single fibers and muscle cross-sections. In the whole homogenates of half of the muscle, cut longitudinally, expression of myosin heavy chain (MHC) will be analyzed. Phosphorylation of the ribosomal protein S6 and 27 kD heat shock protein will be also measured to estimate the rate of protein synthesis. As for the estimation of protein degradation, ubiquitination of MHC will be determined. Further, the number of spermatozoon in testis, which is closely associated with testosterone level, will be also measured, since it was suggested that hindlimb suspension of adult male mice caused a decrease in plasma testosterone, which may play some role(s) in the regulation of muscle mass.
Responses of cell body size and oxidative enzyme activity in motoneurons of the mouse spinal cord following space flight
Ishihara Akihiko, Kyoto University
Tissue: Spinal cord
The major research objective in this study is to compare changes in cell body size and oxidative enzyme activity of motoneurons at the cervical and lumbar segments in the mouse spinal cord following space flight. This study will observe in microgravity a decreased oxidative enzyme activity of motoneurons at the lumbar, but not at the cervical, segment in the mouse spinal cord; no change in cell body size of motoneurons at the cervical or lumbar segment in the mouse spinal cord.
Effects of microgravity on expression and localization of vascular myocyte calcium release channels and endothelial NOS
Morel, Centre National de la Recherche Scientifique, Toulouse, France
In this project we propose to investigate the effects of microgravity on expression and localization of vascular myocyte calcium release channels and endothelial NOS. Cardiovascular deconditioning in microgravity may be mediated through adaptation of postganglionic adrenergic neurons and adrenoreceptors. This process, may in turn be modulated by changes in caloric intake.
Microgravity effects on skeletal muscles Dieter Blottner, Ph.D., Charite Campus Benjamin Franklin, Berlin, Germany
In this project we propose to investigate the NOS /NO pathway expression in mouse hind limb skeletal muscle of the MDS mice and to monitor atrophy and selective proteolysis in skeletal muscle myofibers 1 and 2 of the MDS mice. We expect to confirm that the NOS/NO pathway will be affected (i.e., reduced expression, myofiber type specific expression) by real space flight, and that microgravity-induced muscle atrophy might be monitored by proteolysis biomarkers MuRF1 in slow and fast myofibers of weight bearing or non-weight bearing hind limb muscles in mice. The results may be used to further understand the molecular and cellular adaptation processes to microgravity in vertebrate skeletal muscle. In addition, reliable biomarkers may be helpful to develop effective countermeasure protocols to prevent dramatic muscle atrophy and to ensure performance control for humans in space flight.
Microgravity-induced skin atrophy
Betty Nusgens, Ph.D., University of Liege, Wallonia, Belgium
We have previously demonstrated that fibroblasts from the dermis sense and react to mechanical forces issued from the surrounding extracellular matrix. Conversely, fibroblasts are able to remodel their environment by a dynamic process of synthesis and degradation. Fibroblasts and cells of the vascular network also react to soluble mediators secreted by the keratinocytes such as IL-1, IL-6, VEGF. Skin fibroblasts upon relaxation of mechanical tension adopt a catabolic phenotype and produce significant amount of IL-1 and IL-6, of various matrix metalloproteinases (MMP) while the collagen synthesis is reduced. In microgravity, we have shown that the expression of MMP-1 and IL-6 by dermal fibroblasts is increased indicating that cell might interpret microgravity as a mechanical relaxation. This concept is supported by a reduction of the actin stress fibers and focal adhesions in fibroblasts in microgravity. The expression of IL-1, IL-6 and MMP-1 is under the control of signaling molecules regulating the cytoskeleton dynamics, such as Cdc42, a member of the RhoGTPases family. Another member, Rac1, modulates keratinocytes differentiation and fibroblasts proliferation. Altogether, microgravity might alter metabolic equilibrium of the skin and perhaps of other soft connective tissues such as tendons or interstitial matrix of internal organs.
Astronauts suffer from a significant loss of bone mass during space flight, the ISS Medical Project office has developed some countermeasures to hinder the rapid loss of bone mass. Despite these countermeasures bone mass loss continues to be a problem for astronauts. Finding additional countermeasures will increase the overall health of astronauts on long duration missions.
Microgravity is considered by the scientific community as an accelerated model for studying terrestrial osteoporosis. Results obtained in this space experiment will facilitate the understanding of genetic elements that protect people from osteoporosis. The targeted users are crewmembers after a long-term space mission, elderly people (especially post-menopausal women), and patients after long-time immobilization.
For the MDS experiment, three groups of mice were utilized. One group of mice has been sent to the ISS and was housed in an MDS enclosure. Two control groups of mice remained on Earth in Genoa, Italy; one in a MDS enclosure and the other in standard rodent housing.
On board the ISS, MDS is relatively self-sufficient; the PI team from Genoa checked the health status of the rodents on a daily basis by assessing them through the camera on the viewing window. Water levels were also assessed by the PI team from Genoa daily and refilled by crewmembers as needed. Replacement of the food bars and waste filters were conducted in flight by crewmembers every 20-days or as needed. After landing, the MDS was returned to the investigator for extensive analysis.
Once the rodents were in space, the PI team from Genoa checked the health status of the rodents on a daily basis by assessing them though the camera placed on the viewing window on each MDS. Water levels on the water boxes were checked daily by the PI team from Genoa and refilled by the crewmembers on the ISS as needed. Replacement of the food bars and replacement of the waste filters was conducted in flight by crewmembers every 20 days.
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Mice Drawer System (MDS) reached the ISS on board Shuttle Discovery Flight 17A/STS-128 on August 28, 2009. MDS returned to Earth on November 27, 2009, on Shuttle Atlantis Flight ULF3/STS-129 after a 91-day stay, performing the longest duration of mice in space. MDS flew three wild-type (Wt) and three pleiotrophic transgenic (PTN) mice to determine the microgravity effects levied on each mouse. Unfortunately, during the investigation, one PTN mouse and two Wt mice died due to either health related or payload-related reasons MDS participated in a Tissue Sharing Program with 20 different research groups in order to determine if microgravity induced any tissue modifications, with a primary focus on bone loss. (Cancedda 2012).
Bone Turnover in Wild Type and Pleiotrophin-Transgenic Mice Housed for Three Months in the International Space Station
Ranieri Cancedda, University of Genoa, Italy
Tissue: Bone, Bone Marrow
One of the major goals of the MDS experiment was to investigate bone alterations in 3 Wt and 3 PTN-Tg male mice (2 months old at the time of launch) after 3-month permanence on board the ISS.
This study revealed bone loss during space flight in the weight-bearing bones of both strains. For both PTN-Tg and Wt mice a decrease of the trabecular number as well as an increase of the mean trabecular separation was observed after flight, whereas trabecular thickness did not show any significant change.
Non weight-bearing bones were not affected. The PTN-Tg mice exposed to normal gravity presented a poorer trabecular organization than Wt mice, but interestingly, the expression of the PTN transgene during flight resulted in some protection against microgravity’s’ negative effects. Moreover, osteocytes of the Wt mice, but not of PTN-Tg mice, acquired a round shape, thus showing for the first time osteocyte space-related morphological alterations in vivo. The analysis of specific bone formation and resorption marker expression suggested that the microgravity-induced bone loss was due to both an increased bone resorption and a decreased bone deposition. Apparently, the PTN-Tg protection was the result of a higher osteoblast activity in the flight of mice (Tavella 2012)
Effects of Long-term Space Flight on Erythrocytes and Oxidative Stress of Rodents
Angela Maria Rizzo, University of Milan, Milan, Italy
Several hematological modifications in humans were observed after microgravity exposure such as depression of T-cell lymphocyte activation and of our ability to fight infectious microorganisms. Moreover erythrocyte (red blood cell) hemolysis (breakdown) and hemoglobin loss was also observed after space missions. In addition space radiations can induce generation of hydroxyl radicals, very reactive at the site of their formation, which can initiate a chain of reactions leading to lipid peroxidation (causing cell damage).
In the MDS mice, after landing, blood cell parameter showed a higher erythrocyte concentration (RBC) and RDW% with a hematocrit near or above 50%. Platelets were increased in both Wt and TG mice after flight, while hemoglobin content remained constant. These data are partially in accordance with human data after space mission and could probably be the consequence of body fluid shift and altered renal function.
Both Wt and Tg mice after space flight underwent to an oxidative stress and an increase of by-products of lipid peroxidation concentration. In parallel antioxidant and enzymes involved in the elimination of hydroperoxides from lipids were also enhanced with higher levels in Tg.
After the MDS mission, mice erythrocytes presented modifications in the cell membrane composition and an increase of lipid peroxidation products. Despite their cellular activation, antioxidant defences were not sufficient to prevent damages caused by oxidative stresses (Rizzo 2012).
Adaptation of mouse skeletal muscle to long-term microgravity in the MDS mission
Stefano Schiaffino, University of Padova, Padua, Italy
Diana Conte Camerino University of Bari, Bari, Italy
Tissue: Limb Skeletal Muscles
So far, the effect of microgravity on skeletal muscles has been examined in mice only after a short-term (5–20 day) exposition. It has been observed that space flight has adverse effects on muscles, including atrophy and partial shift of muscle fibres toward a faster, more glycolytic phenotype.
The MDS experiment gave the first opportunity to study long time exposure to microgravity effects on skeletal muscles. After the 91 day flight, muscle atrophy was observed in the fibres of the soleus muscle, but this atrophy was only slightly increased when compared to shorter period in microgravity. Alterations were observed in the soleus and to a lesser extent in the extensor digitorum longus (EDL) muscle, in particular with regard to slow-to-fast fibre transition and ion channel activity.
Gene expression of the atrophy-related ubiquitin ligases was up-regulated in both soleus and EDL muscles from flight mice, whereas autophagy (self-degradation) was in the control range. In the same animals, various stress-related genes were up-regulated in EDL, but not in soleus. On the all, gene expression results suggested that EDL muscle may resist to microgravity-induced atrophy by activating compensatory and protective mechanisms and identified some molecular targets for the development of countermeasures (Sandonà 2012).
The Impact of Long-Term Exposure to Space Environment on Adult Mammalian Organisms: A Study on a Mouse Thyroid and Testis
Human hormonal levels are known to change during space flight, but the underlying mechanisms are still unknown. To clarify this point, thyroid and testis/epididymis from the flight and control mice were analysed both morphologically and functionally.
While Wt ground samples showed variable size and spatial orientation, space flight animals had a more homogenous thyroid tissue structure with a reduction in the interior spacing. In space flight animals both in Wt and Tg mice the follicular size was greatly varied.
Structural modification correlated with altered thyroid functionality. Both Wt and Tg cells stimulated with thyrotropin enhanced cAMP production in ground cells. After space flight it more pronounced enhancement with Wt mice was observed. At the end of the MDS experiment, the thyrotropin receptor and caveolin-1 in the thyroid were overexpressed in the space flight mice.
In testes, the space flight mice showed severe degenerative changes, in some cases with tubules almost devoid of spermatozoa except for few spermatogonia. Tubular degeneration was not homogeneous and differences were not seen between Wt and Tg mice. The expression of androgen and follicle stimulating hormone receptors increased while luteinizing hormone receptor levels were not changed.
These data indicate that several changes occur in relevant endocrine organs under the control of the pituitary gland and they could be responsible for variations of hormone levels in human during space missions, significantly affecting the endocrine homeostasis of the body, as well the reproductive function (Masini 2012).
Evaluation of Gene, Protein and Neurotrophin Expression in the Brain of Mice Exposed to Space Environment for 91 Days
Daniela Santucci, Istituto Superiore di Sanita’, Rome, Italy
Yoshinobu Ohira, Osaka University, Osaka, Japan
Tissue: Brain, Adrenal Glands, Tongue, Spinal Cord
While modification in the central nervous system are described in literature after short space mission, long-term inhibition of antigravity activity on mouse brain are unclear. After the MDS experiment, the effects of 3-month exposure to microgravity environment on the expression of genes and proteins in mouse brain were studied.
Several genes related to the immune response, metabolic process, and/or inflammatory responses were up-regulated whereas several genes involved in various metabolic and catabolic processes were down-regulated. Two proteins, BDNF and NGF, which are involved in learning and memory performance, ageing-related disorders and anxiety-like behaviour were studied in brain and adrenal gland. Expression of NGF in hippocampus, cortex, and adrenal gland of wild type animals tended to decrease following space flight, but together with BDNF it was not consistent suggesting only a transient response to space flight and not long-lasting effects. On the contrary CRMP1 was up-regulated in flight samples and, since the CRMP1 deficient mice showed an impaired spatial learning and memory, memory performance may be stimulated after space flight.
Exposure to space environment influenced the expression of a number of genes and proteins in the brain that have been shown to be involved in a wide spectrum of biological function and appears to interfere with expression of neuropeptides involved in psycho-neuro-endocrine adaptations that should need a deeper examination in a MDS re-flight (Masini 2012).
Loss of Parafollicular Cells during Gravitational Changes (Microgravity, Hypergravity) and the Secret Effect of Pleiotrophin
Bone loss is one of the most important complications for crewmembers who are exposed to long-term microgravity. Changes in blood flow and systemic hormones were indicated as important contributing elements to the response of the mechanical loading experienced by osteoblast cells. Here, the possible biological involvement of thyroid C cells is being investigated.
This study has provided evidence that both microgravity and hypergravity induce similar loss of thyroid C cells with reduction of calcitonin production. Pleiotrophin over-expression results in some protection against negative effects of gravity change. To confirm these results it would be important to know blood levels of calcitonin in hypogravity and hypergravity environments and this could be an area of study for future missions (Albi[a] 2012).
Observing the Mouse Thyroid Sphingomyelin Under Space Conditions: A Case Study from the MDS Mission in Comparison with Hypergravity Conditions
Histological examination of the thyroid gland revealed an increase in the average follicle size compared to that of three control animals and three animals exposed to hypergravity (2g) conditions in a centrifuge. Additional analysis detected an increase in two thyroid gland enzymes, sphingomyelinase and sphingomyelin-synthase1. In addition, sphingomyelinase, an enzyme traditionally confined to the cell nucleus in the control animals, was found in the mouse exposed to hypogravity to be homogenously distributed throughout the cell bodies (Albi[b] 2012).
Evaluation of long term space permanence effects on other tissues of mice exposed to space environment for 3 months
Additional tissues are currently being processed and additional data collected by the different PIs.
Masini MA, Albi E, Barmo C, Bonfiglio T, Bruni L, Canesi L, Cataldi S, Curcio F, D'Amora M, Ferri I, Goto K, Kawano F, Lazzarini R, Loreti E, Nakai N, Ohira T, Ohira Y, Palmero S, Prato P, Ricci F, Scarabelli L, Shibaguchi T, Spelat R, Strollo F, Ambesi-Impiombato FS. The Impact of Long-Term Exposure to Space Environment on Adult Mammalian Organisms: A Study on Mouse Thyroid and Testis. PLOS ONE. 2012; 7(4): e35418. DOI: 10.1371/journal.pone.0035418.
Albi E, Curcio F, Lazzarini A, Floridi A, Cataldi S, Lazzarini R, Loreti E, Ferri I, Ambesi-Impiombato FS. How microgravity changes Galectin-3 in thyroid follicles. BioMed Research International. 2014; 2014(652863): 5 pp. DOI: 10.1155/2014/652863.
Albi E, Curcio F, Spelat R, Lazzarini A, Lazzarini R, Loreti E, Ferri I, Ambesi-Impiombato FS. Observing the Mouse Thyroid Sphingomyelin Under Space Conditions: A Case Study from the MDS Mission in Comparison with Hypergravity Conditions. Astrobiology. 2012 October 19; 12(11): 1035-1041. DOI: 10.1089/ast.2012.0881. PMID: 23082746. [b]
Cancedda R, Liu Y, Ruggiu A, Tavella S, Biticchi R, Santucci D, Schwartz S, Ciparelli P, Falcetti G, Tenconi C, Cotronei V, Pignataro S. The Mice Drawer System (MDS) Experiment and the Space Endurance Record-Breaking Mice. PLOS ONE. 2012; 7(5): e32243. DOI: 10.1371/journal.pone.0032243.
Santucci D, Kawano F, Ohira T, Terada M, Nakai N, Francia N, Alleva E, Aloe L, Ochiai T, Cancedda R, Goto K, Ohira Y. Evaluation of Gene, Protein and Neurotrophin Expression in the Brain of Mice Exposed to Space Environment for 91 Days. PLOS ONE. 2012; 7(7): e40112. DOI: 10.1371/journal.pone.0040112.
Rizzo AM, Corsetto PA, Montorfano G, Milani S, Zava S, Tavella S, Cancedda R, Berra B. Effects of Long-Term Space Flight on Erythrocytes and Oxidative Stress of Rodents. PLOS ONE. 2012; 7(3): e32361. DOI: 10.1371/journal.pone.0032361.
Neutelings T, Nusgens B, Liu Y, Tavella S, Ruggiu A, Cancedda R, Gabriel M, Colige A, Lambert C. Skin physiology in microgravity: a 3-month stay aboard ISS induces dermal atrophy and affects cutaneous muscle and hair follicles cycling in mice. npj Microgravity. 2015 May 27; 1: 15002. DOI: 10.1038/npjmgrav.2015.2.
Tavella S, Ruggiu A, Guiliana A, Brun F, Canciani B, Manescu A, Marozzi K, Cilli M, Costa D, Liu Y, Piccardi F, Tasso R, Tromba G, Rustichelli F, Cancedda R. Bone Turnover in Wild Type and Pleiotrophin-Transgenic Mice Housed for Three Months in the International Space Station (ISS). PLOS ONE. 2012; 7(9): e33179. DOI: 10.1371/journal.pone.0033179.
Camerino GM, Pierno S, Liantonio A, De Bellis M, Cannone M, Sblendorio V, Conte E, Mele A, Tricarico D, Tavella S, Ruggiu A, Cancedda R, Ohira Y, Danieli-Betto D, Ciciliot S, Germinario E, Sandona D, Betto R, Camerino DC, Desaphy J. Effects of Pleiotrophin Overexpression on Mouse Skeletal Muscles in Normal Loading and in Actual and Simulated Microgravity. PLOS ONE. 2013 August 28; 8(8): e72028. DOI: 10.1371/journal.pone.0072028.
Albi E, Curcio F, Spelat R, Lazzarini A, Lazzarini R, Cataldi S, Loreti E, Ferri I, Ambesi-Impiombato FS. Loss of Parafollicular Cells during Gravitational Changes (Microgravity, Hypergravity) and the Secret Effect of Pleiotrophin. PLOS ONE. 2012; 7(12): e48518. DOI: 10.1371/journal.pone.0048518. PMID: 23284618. [a]
Ohira T, Ohira T, Kawano T, Shibaguchi T, Okabe H, Goto K, Ogita F, Sudoh M, Roy RR, Edgerton VR, Cancedda R, Ohira Y. Effects of gravitational loading levels on protein expression related to metabolic and/or morphologic properties of mouse neck muscles. Physiological Reports. 2014 January; 2(1). DOI: 10.1002/phy2.183.
Sandona D, Desaphy J, Camerino GM, Bianchini E, Ciciliot S, Danieli-Betto D, Dobrowolny G, Furlan S, Germinario E, Goto K, Gutsmann M, Kawano F, Nakai N, Ohira Y, Ohno Y, Picard A, Salanova M, Schiffl G, Blottner D, Musaro A, Betto R, Camerino DC, Schiaffino S. Adaptation of Mouse Skeletal Muscle to Long-Term Microgravity in the MDS Mission. PLOS Biology. 2012 Mar 28; 7(3): e33232. DOI: 10.1371/journal.pone.0033232.
Ground Based Results Publications
Albi E, Ambesi-Impiombato FS, Lazzarini A, Lazzarini R, Floridi A, Cataldi S, Loreti E, Ferri I, Curcio F. Reinterpretation of mouse thyroid changes under space conditions: The contribution of confinement to damage. Astrobiology. 2014 July; 14(7): 563-567. DOI: 10.1089/ast.2014.1166. PMID: 24945896.
Cancedda R, Pignataro S, Alberici G, Tenconi C. Mice Drawer System: phase c/d development and perspective. Journal of Gravitational Physiology. 2002; 9(1): P337-338. PMID: 15002603.
Italian Mice to Head Into Space
MDS integrated inside the Double Payload Container. Image provided courtesy of ASI.
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Lateral side view of MDS, with waste filter partially removed. Image courtesy of ASI.
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Side view: facility includes cages that will house six individual mice. Image courtesy of ASI.
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NASA Image: ISS020E049908: NASA astronaut Nicole Stott, Expedition 20/21 flight engineer, is pictured near the Mice Drawer System (MDS) in the Kibo laboratory of the International Space Station.
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NASA Image: ISS020E049909 - NASA astronaut Nicole Stott, Expedition 20/21 flight engineer, works with the Mice Drawer System (MDS) in the Kibo laboratory of the International Space Station.
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NASA Image: S128E007107 - Astronauts Nicole Stott, Expedition 20 flight engineer; and Patrick Forrester, STS-128 mission specialist, work in the Kibo laboratory of the International Space Station while Space Shuttle Discovery remains docked to the station.
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