Fumiaki Tanigaki

Microgravity promotes osteoclast activity in medaka fish reared at the international space station

The bone mineral density (BMD) of astronauts decreases specifically in the weight-bearing sites during spaceflight. It seems that osteoclasts would be affected by a change in gravity; however, the molecular mechanism involved remains unclear. Here, we show that the mineral density of the pharyngeal bone and teeth region of TRAP-GFP/Osterix-DsRed double transgenic medaka fish was decreased and that osteoclasts were activated when the fish were reared for 56 days at the international space station. In addition, electron microscopy observation revealed a low degree of roundness of mitochondria in osteoclasts. In the whole transcriptome analysis, fkbp5 and ddit4 genes were strongly up-regulated in the flight group. The fish were filmed for abnormal behavior; and, interestingly, the medaka tended to become motionless in the late stage of exposure. These results reveal impaired physiological function with a change in mechanical force under microgravity, which impairment was accompanied by osteoclast activation.

Four-year bacterial monitoring in the International Space Station—Japanese Experiment Module “Kibo” with culture-independent approach

Studies on the relationships between humans and microbes in space habitation environments are critical for success in long-duration space missions, to reduce potential hazards to the crew and the spacecraft infrastructure. We performed microbial monitoring in the Japanese Experiment Module “Kibo”, a part of the International Space Station, for 4 years after its completion, and analyzed samples with modern molecular microbiological techniques. Sampling was performed in September 2009, February 2011, and October 2012. The surface of the incubator, inside the door of the incubator, an air intake, air diffuser, and handrail were selected as sampling sites. Sampling was performed using the optimized swabbing method. Abundance and phylogenetic affiliation of bacteria on the interior surfaces of Kibo were determined by quantitative PCR and pyrosequencing, respectively. Bacteria in the phyla Proteobacteria (γ-subclass) and Firmicutes were frequently detected on the interior surfaces in Kibo. Families Staphylococcaceae and Enterobacteriaceae were dominant. Most bacteria detected belonged to the human microbiota; thus, we suggest that bacterial cells are transferred to the surfaces in Kibo from the astronauts. Environmental bacteria such as Legionella spp. were also detected. From the data on bacterial abundance and phylogenetic affiliation, Kibo has been microbiologically well maintained; however, the microbial community structure in Kibo may change with prolonged stay of astronauts. Continuous monitoring is required to obtain information on changes in the microbial community structure in Kibo.

Space Habitation and Microbiology: Status and Roadmap of Space Agencies

The ubiquitous nature of microbiology is reflected in the diversity of microbiological research and operational efforts at NASA. For example, the impact of microorganisms on other planets and the protection of Earth from the potential of microbial life elsewhere is the responsibility of the Office of Planetary Protection (http://planetaryprotection.nasa.gov/ about). While the Office of Planetary Protection does not include forward or back contamination to or from a low Earth orbit, research platforms, such as the ISS, are being used to better understand the survival of microorganisms and corresponding contamination control in the extreme conditions of space (2, 4, 10). Another example is the NASA Center for Astrobiology, which focuses on the origin, evolution, distribution, and future of life in the universe (https:// astrobiology.nasa.gov/). The large focus on microbiology is within its human exploration operations. NASA has historically set microbiological requirements, including stringent monitoring regimes, to mitigate risks to the health and performance of astronauts. Microorganisms can have both positive and negative impacts on many aspects of human spaceflight, including the risk and prevention of infectious diseases, performance of Environmental Control and Life Support Systems (ECLSS), spaceflight foods, and vehicle design and integrity. Even though a great amount of information has been obtained, several key questions regarding the impact of microorganisms on human spaceflight still remain. Research into the uncertainties of risks that may affect crew health are the responsibility of the NASA Human Research Program (http://www.nasa.gov/exploration/human research/). The research that addresses our fundamental understanding of space life science and its translation for benefits to the general public on Earth is the responsibility of NASA Space Biology.

Acute transcriptional up-regulation specific to osteoblasts/osteoclasts in medaka fish immediately after exposure to microgravity

Bone loss is a serious problem in spaceflight; however, the initial action of microgravity has not been identified. To examine this action, we performed live-imaging of animals during a space mission followed by transcriptome analysis using medaka transgenic lines expressing osteoblast and osteoclast-specific promoter-driven GFP and DsRed. In live-imaging for osteoblasts, the intensity of osterix- or osteocalcin-DsRed fluorescence in pharyngeal bones was significantly enhanced 1 day after launch; and this enhancement continued for 8 or 5 days. In osteoclasts, the signals of TRAP-GFP and MMP9-DsRed were highly increased at days 4 and 6 after launch in flight. HiSeq from pharyngeal bones of juvenile fish at day 2 after launch showed up-regulation of 2 osteoblast- and 3 osteoclast- related genes. Gene ontology analysis for the whole-body showed that transcription of genes in the category “nucleus” was significantly enhanced; particularly, transcription-regulators were more up-regulated at day 2 than at day 6. Lastly, we identified 5 genes, c-fos, jun-B-like, pai-1, ddit4 and tsc22d3, which were up-regulated commonly in the whole-body at days 2 and 6, and in the pharyngeal bone at day 2. Our results suggested that exposure to microgravity immediately induced dynamic alteration of gene expression levels in osteoblasts and osteoclasts.

Histological and Transcriptomic Analysis of Adult Japanese Medaka Sampled Onboard the International Space Station

To understand how humans adapt to the space environment, many experiments can be conducted on astronauts as they work aboard the Space Shuttle or the International Space Station (ISS). We also need animal experiments that can apply to human models and help prevent or solve the health issues we face in space travel. The Japanese medaka (Oryzias latipes) is a suitable model fish for studying space adaptation as evidenced by adults of the species having mated successfully in space during 15 days of flight during the second International Microgravity Laboratory mission in 1994. The eggs laid by the fish developed normally and hatched as juveniles in space. In 2012, another space experiment (“Medaka Osteoclast”) was conducted. Six-week-old male and female Japanese medaka (Cab strain osteoblast transgenic fish) were maintained in the Aquatic Habitat system for two months in the ISS. Fish of the same strain and age were used as the ground controls. Six fish were fixed with paraformaldehyde or kept in RNA stabilization reagent (n = 4) and dissected for tissue sampling after being returned to the ground, so that several principal investigators working on the project could share samples. Histology indicated no significant changes except in the ovary. However, the RNA-seq analysis of 5345 genes from six tissues revealed highly tissue-specific space responsiveness after a two-month stay in the ISS. Similar responsiveness was observed among the brain and eye, ovary and testis, and the liver and intestine. Among these six tissues, the intestine showed the highest space response with 10 genes categorized as oxidation–reduction processes (gene ontogeny term GO:0055114), and the expression levels of choriogenin precursor genes were suppressed in the ovary. Eleven genes including klf9, klf13, odc1, hsp70 and hif3a were upregulated in more than four of the tissues examined, thus suggesting common immunoregulatory and stress responses during space adaptation.