Physical determinants of radiation sensitivity in bacterial spores.

Several factors modifying radiation sensitivity in dry bacterial spores are described and discussed. Vacuum inducing the loss of critical structural water, very low dose rates of radiation from which the cell may recover, radiations of high linear energy transfer, and the action of temperature over long periods of time on previously irradiated cells are recognized from extensive laboratory work as important in determining survival of spores exposed to low radiation doses at low temperatures for long periods of time. Some extensions of laboratory work are proposed.     

Long-term space flights--personal impressions.

During a final 4-month stage of a 1-year space flight of cosmonauts Titov and Manarov, a physician, Valery Polyakov was included on a crew for the purpose of evaluating their health, correcting physical status to prepare for the spacecraft reentry and landing operations. The complex program of scientific investigations and experiments performed by the physician included an evaluation of adaptation reactions of the human body at different stages of space mission using clinicophysiological and biochemical methods; testing of alternative regimes of exercise and new countermeasures to prevent an unfavourable effect of long-term weightlessness.     

Viability of bacterial spores exposed to hydrazine

For the purposes of planetary protection, a series of experiments were performed to answer a long-standing question about the potential of bacterial contamination of interplanetary spacecraft from liquid hydrazine. Spores of Bacillus atrophaeus (ATCC No. 9372, also known as Bacillus subtilis var. niger, and BSN) were exposed to hydrazine and survivors were enumerated using the NASA standard planetary protection pour plate assay. Results indicate that bulk hydrazine rocket propellant may be considered free of living bacterial cells for planetary protection compliance.     

Differentiation of Dictyostelium discoideum vegetative cells into spores during Earth orbit in space.

We reported previously that emerged amoebae of Dictyostelium (D.) discoideum grew, aggregated and differentiated to fruiting bodies with normal morphology in space. Here, we investigated the effects of space radiation and/or microgravity on the number, viability, kinetics of germination, growth rate and mutation frequency of spores formed in space in a radiation-sensitive strain, gamma s13, and the parental strain, NC4. In gamma s13, there were hardly spores in the fruiting bodies formed in space. In NC4, we found a decrease in the number of spores, a delay in germination of the spores and delayed start of cell growth of the spores formed in space when compared to the ground control. However, the mutation frequency of the NC4 spores formed in space was similar to that of the ground control. We conclude that the depression of spore formation might be induced by microgravity and/or space radiation through the depression of some stage(s) of DNA repair during cell differentiation in the slime mold.     

Long-term modulation of Galactic Cosmic Radiation and its model for space exploration.

As the human exploration of space has received new attention in the United States, studies find that exposure to space radiation could adversely impact the mission design. Galactic Cosmic Radiation (GCR), with its very wide range of charges and energies, is particularly important for a mission to Mars, because it imposes a stiff mass penalty for spacecraft shielding. Dose equivalent versus shielding thickness calculations, show a rapid initial drop in exposure with thickness, but an asymptotic behavior at a higher shielding thickness. Uncertainties in the radiobiology are largely unknown. For a fixed radiation risk, this leads to large uncertain ties in shielding thickness for small uncertainties in estimated dose. In this paper we investigate the application of steady-state, spherically-symmetric diffusion-convection theory of solar modulation to individual measurements of differential energy spectra from 1954 to 1989 in order to estimate the diffusion coefficient, kappa (r,t), as a function of time. We have correlated the diffusion coefficient to the Climax neutron monitor rates and show that, if the diffusion coefficient can be separated into independent functions of space and time: kappa (-r,t)=K(t)kappa 0 beta P kappa 1(r), where beta is the particle velocity and P the rigidity, then (i) The time dependent quantity 1/K(t), which is proportional to the deceleration potential, phi(r,t), is linearly related to the Climax neutron monitor counting rate. (ii) The coefficients obtained from hydrogen or helium intensity measurements are the same. (iii) There are different correlation functions for odd and even solar cycles. (iv) The correlation function for the Climax neutron monitor counting rate for given time, t, can be used to estimate mean deceleration parameter phi(t) to within +/- 15% with 90% confidence. We have shown that kappa(r,t) determined from hydrogen and/or helium data, can be used to fit the oxygen and iron differential energy spectra with a root mean square error of about +/- 10%, and essentially independent of the particle charge or energy. We have also examined the ion chamber and 14C measurements which allow the analysis to be extended from the year 1906 to 1990. Using this model we have defined reference GCR spectra at solar minimum and solar maximum. These can be used for space exploration studies and provide a quantitative estimate of the error in dose due to changes in GCR intensities.     

Oxygen effect, hydrogen peroxide yields, and time scale of interaction of potentially damaging species in electron pulse irradiated bacterial spores.

A given integrated radiation dose delivered from a LINAC as a train of pulses (50/s), characteristically of 0.1 to 5 microseconds pulse length with dose rates within the pulse between 0.38 and 38 krads/microsecond, inactivates bacterial spores in water suspension more effectively than the same dose given as Co60 gamma rays. This enhancement of radiation damage occurs both in the presence and in the absence of oxygen and is not explained by either pulse dose rate or pulse length alone, but is monotonically related to the product of these pulse parameters, pulse dose. The enhancement appears to result from the interaction, within individual spores, of free radical species of average lifetime of about 2-5 microseconds. The time scale over which these species operate suggests that they are freely diffusable. Prevention, in part, of their damaging effect by the presence of selective scavenging agents is evidence that OH radicals are involved. Measurements of H2O2 yield for irradiation conditions that show a gradation of enhancement of damage correlate strongly with the extent of damage observed.     

Fullerenes in space.

The discovery and synthesis of fullerenes led to the hypothesis that they may be present and stable in interstellar space. Fullerenes have been reported in an impact crater on the LDEF spacecraft. Investigations of fullerenes in carbonaceous meteorites have yielded only small upper limits. Fullerene compounds and their ions could be interesting carrier molecules for some of the "diffuse interstellar bands" (DIBs), a long standing mystery in astronomy. We have detected two new diffuse bands that are consistent with laboratory measurements of the C60+, as first evidence for the largest molecule ever detected in space. Criteria for this identification are discussed. The inferred abundance (up to 0.9 % of cosmic carbon locked in C60+) suggests that fullerenes may play an important role in interstellar chemistry. We present new observations on DIB substructures consistent with fullerene compounds, and the search for neutral C60 in the diffuse medium.     

Pituitary cells in space.

Cells of the mammalian pituitary gland synthesize and secrete several protein hormones which regulate a number of organ systems throughout the body. These include the musculoskeletal, immune, vascular and endocrine systems. Since changes occur in these tissues as a result of spaceflight, and since pituitary growth hormone (GH) and prolactin (PRL) play a role in the control of these systems on earth, we have focused attention over the last 10 years on GH and PRL cell function during and after spaceflight. The cumulative results of 4 spaceflight missions and several mimicked microgravity experiments establish 1) that production and release of biologically active GH and PRL is repeatedly and significantly attenuated (usually > 50%) and 2) that changes in cell morphology also occur. In this paper we describe our results within the framework of methodologies and approaches frequently used to study pituitary cell function on earth. In so doing we hope to develop future flight experiments aimed at uncovering possible microgravity "sensing systems" within the pituitary cell.     

Preliminary results of space experiment: Implications for the effects of space radiation and microgravity on survival and mutation induction in human cells

In view of the concern for the health of astronauts that may one day journey to Mars or the Moon, we investigated the effect that space radiation and microgravity might have on DNA damage and repair. We sent frozen human lymphoblastoid TK6 cells to the International Space Station where they were maintained under frozen conditions during a 134-day mission (14 November 2008 to 28 March 2009) except for an incubation period of 8 days under 1G or μG conditions in a CO₂ incubator. The incubation period started after 100 days during which the cells had been exposed to 54 mSv of space radiation. The incubated cells were then refrozen, returned to Earth, and compared to ground control samples for the determination of the influence of microgravity on cell survival and mutation induction. The results for both varied from experiment to experiment, yielding a large SD, but the μG sample results differed significantly from the 1G sample results for each of 2 experiments, with the mean ratio of μG to 1G being 0.55 for the concentration of viable cells and 0.59 for the fraction of thymidine kinase deficient (TK⁻) mutants. Among the mutants, non-loss of zygosity events (point mutations) were less frequent (31%) after μG incubation than after 1G incubation, which might be explained by the influence of μG on cellular metabolic or physiological function. Additional experiments are needed to clarify the effect of μG interferes on DNA repair.     

Mammalian cell cultivation in space.

Equipment used in space for the cultivation of mammalian cells does not meet the usual standard of earth bound bioreactors. Thus, the development of a space worthy bioreactor is mandatory for two reasons: First, to investigate the effect on single cells of the space environment in general and microgravity conditions in particular, and second, to provide researchers on long term missions and the Space Station with cell material. However, expertise for this venture is not at hand. A small and simple device for animal cell culture experiments aboard Spacelab (Dynamic Cell Culture System; DCCS) was developed. It provides 2 cell culture chambers, one is operated as a batch system, the other one as a perfusion system. The cell chambers have a volume of 200 microliters. Medium exchange is achieved with an automatic osmotic pump. The system is neither mechanically stirred nor equipped with sensors. Oxygen for cell growth is provided by a gas chamber that is adjacent to the cell chambers. The oxygen gradient produced by the growing cells serves to maintain the oxygen influx by diffusion. Hamster kidney cells growing on microcarriers were used to test the biological performance of the DCCS. On ground tests suggest that this system is feasible.     

Human reproductive issues in space.

Animal studies in space or analogous environments have suggested that there may be problems in the reproductive sphere; such factors might limit mankind's ability to live and work for extended periods of time in microgravity or on non-terrestrial planetary surfaces. A review of reproductive functioning in animal species studied during space flight demonstrated that most species were affected significantly by the absence of gravity and/or the presence of radiation. These two factors induced alterations in normal reproductive functioning independently of, as well as in combination with, each other. Based on animal models, we have identified several potential problem areas regarding human reproductive physiology and functioning in the space environment. While there are no current space flight investigations, the animal studies suggest priorities for future research in human reproduction. Such studies will be critical for the successful colonization of the space frontier.     

Thin film bioreactors in space.

Studies from the Skylab, SL-3 and D-1 missions have demonstrated that biological organisms grown in microgravity have changes in basic cellular functions such as DNA, mRNA and protein synthesis, cytoskeleton synthesis, glucose utilization and cellular differentiation. Since microgravity could affect prokaryotic and eukaryotic cells at a subcellular and molecular level, space offers us an opportunity to learn more about basic biological systems with one important variable removed. The thin film bioreactor will facilitate the handling of fluids in microgravity, under constant temperature and will allow multiple samples of cells to be grown with variable conditions. Studies on cell cultures grown in microgravity would enable us to identify and quantify changes in basic biological function in microgravity which are needed to develop new applications of orbital research and future biotechnology.     

Progress in plant research in space.

Progress is reviewed of spaceflight research conducted with plants between 1987 and 1992. Numerous plant experiments have been performed on spacecraft and sounding rockets in the past five years by scientists of the US, the former Soviet Union, Europe, and other areas. The experiments are categorized into three areas: gravity sensing, transduction, and response; development and reproduction; and metabolism, photosynthesis, and transport. The results of these experiments continue to demonstrate that gravity and/or other factors of spaceflight affect plants at the organismal, cellular, subcellular, and molecular levels, resulting in changes in orientation, development, metabolism, and growth. The challenge now is to truly dissect the effects of gravity from those of other spaceflight factors and to identify the basic mechanisms underlying gravity's effects.     

Long-term preservation of microbial ecosystems in permafrost.

It has been established that significant numbers (up to 10 million cells per gram of sample) of living microorganisms of various ecological and morphological groups have been preserved under permafrost conditions, at temperatures ranging from -9 to -13 degrees C and depths of up to 100 m, for thousands and sometimes millions of years. Preserved since the formation of permafrost in sand-clay sediments of the Pliocene-Quaternary period and in paleosols and peats buried among them, these cells art the only living organisms that have survived for a geologically significant period of time. The complexity of the microbial community preserved varies with the age of the permafrost. Eukaryotes are found only in Holocene sediments; while prokaryotes are found to greater ages, i.e., Pliocene and Pleistocene. The diversity of microorganisms decreases with increasing age of sediments, and as a result cocci and corynebacteria are predominant. Enzyme activity (catalase and hydrolytic enzymes) and photosynthetic pigments (chlorophyll and pheophytin have also been detected in permafrost sediments. These results permit us to outline some approaches to the search for traces of life in the permafrost of Martian sediments by borehole core sampling. It is in the deep horizons (and not on the planet surface), isolated by permafrost from the external conditions, that results similar to those obtained on Earth can be expected.     

Production and action of cytokines in space.

B6MP102 cells, a continuously cultured murine bone marrow macrophage cell line, were tested for secretion of tumor necrosis factor-alpha and Interleukin-1 during space flight. We found that B6MP102 cells secreted more tumor necrosis factor-alpha and interleukin-1 when stimulated in space with lipopolysaccharide than controls similarly stimulated on earth. This compared to increased secretion of interferon-beta and -gamma by lymphocytes that was measured on the same shuttle flights. Although space flight enhanced B6MP102 secretion of tumor necrosis factor-alpha, an experiment on a subsequent space flight (STS-50) found that cellular cytotoxicity, mediated by tumor necrosis factor-alpha, was inhibited.     

The minimal cost of life in space.

The cost of keeping people alive in space is assessed from a theoretical viewpoint and using two actual designs for plant growth systems. While life support is theoretically not very demanding, our ability to implement life support is well below theoretical limits. A theoretical limit has been calculated from requirements and the state of the art for plant growth has been calculated using data from the BIO-Plex PDR and from the Cornell CEA prototype system. The very low efficiency of our current approaches results in a high mission impact, though we can still see how to get a significant reduction in cost of food when compared to supplying it from Earth. Seeing the distribution of costs should allow us to improve our current designs.     

Results of dosimetric measurements in space missions.

Detector packages consisting of plastic nuclear track detectors, nuclear emulsions, and theromoluminescence detectors were exposed at different locations inside the space laboratory Spacelab and at the astronauts' body and in different sections of the MIR space station. Total dose, particle fluence rate and linear energy transfer (LET) spectra of heavy ions, number of nuclear disintegrations and fast neutron fluence rates were determined of each exposure. The dose equivalent received by the Payload specialists (PSs) were calculated from the measurements, they range from 190 microSv d-1 to 770 microSv d-1. Finally, a preliminary investigation of results from a particle telescope of two silicon detectors, first used in the last BIORACK mission on STS 76, is reported.     

On the radiosensitivity of man in space.

Astronauts' radiation exposure limits are based on experimental and epidemiological data obtained on Earth. It is assumed that radiation sensitivity remains the same in the extraterrestrial space. However, human radiosensitivity is dependent upon the response of the hematopoietic tissue to the radiation insult. It is well known that the immune system is affected by microgravity. We have developed a mathematical model of radiation-induced myelopoiesis which includes the effect of microgravity on bone marrow kinetics. It is assumed that cellular radiosensitivity is not modified by the space environment, but repopulation rates of stem and stromal cells are reduced as a function of time in weightlessness. A realistic model of the space radiation environment, including the HZE component, is used to simulate the radiation damage. A dedicated computer code was written and applied to solar particle events and to the mission to Mars. The results suggest that altered myelopoiesis and lymphopoiesis in microgravity might increase human radiosensitivity in space.     

Longevity in space; experiment on the life span of Paramecium cell clone in space.

Life span is the most interesting and also the most important biologically relevant time to be investigated on the space station. As a model experiment, we proposed an investigation to assess the life span of clone generation of the ciliate Paramecium. In space, clone generation will be artificially started by conjugation or autogamy, and the life span of the cell populations in different gravitational fields (microgravity and onboard 1 x g control) will be precisely assessed in terms of fission age as compared with the clock time. In order to perform the space experiment including long-lasting culture and continuous measurement of cell division, we tested the methods of cell culture and of cell-density measurement, which will be available in closed environments under microgravity. The basic design of experimental hardware and a preliminary result of the cultivation procedure are described.