[TOLL-LIKE RECEPTORS IN COSMONAUT'S PERIPHERAL BLOOD CELLS AFTER LONG-DURATION MISSIONS TO THE INTERNATIONAL SPACE STATION].

Studies of Toll-like receptors (TLR) in 20 cosmonauts-members of long-duration (124-199-day) missions to the International space station evidenced changes in relative and absolute counts of peripheral blood monocytes with TLR2, TLR4 and TLR6 on the surface, expression of TLR2 and TLR6 genes, and genes of molecules involved in the TLR signaling pathway and TLR-related NF-KB-, JNK/p38- and IRF pathways on the day of return to Earth. The observed changes displayed individual variability.

520-d Isolation and confinement simulating a flight to Mars reveals heightened immune responses and alterations of leukocyte phenotype.

During interplanetary exploration, chronic stress caused by long term isolation and confinement in the spacecraft is one of the major concerns of physical and psychological health of space travelers. And for human on Earth, more and more people live in an isolated condition, which has become a common social problem in modern western society. Collective evidences have indicated prolonged chronic stress could bring big influence to human immune function, which may lead to a variety of health problems. However, to what extent long-term isolation can affect the immune system still remains largely unknow. A simulated 520-d Mars mission provided an extraordinary chance to study the effect of prolonged isolation. Six healthy males participated in this mission and their active neuroendocrine and immune conditions were studied with saliva and blood samples from all participants on chosen time points during the isolation period. As a typical neuroendocrine parameter, stress hormone cortisol was measured in the morning saliva samples. Immune phenotype changes were monitored through peripheral leukocyte phenotype analysis. Using an ex vivo viral infection simulation assay we assessed the immune response changes characterized by the ability to produce representative endogenous pro-inflammatory cytokines. The results of this study revealed elevated cortisol levels, increased lymphocyte amount and heightened immune responses, suggesting that prolonged isolation acting as chronic stressors are able to trigger leukocyte phenotype changes and poorly controlled immune responses.

[Status of the osteoclast-activating system in cosmonauts after long-duration missions to the International Space Station].

The results of studying the system of osteoprotegerin/ receptor activator of nuclear factor kappa-B ligand (OPG/RANKL) in 22 cosmonauts after long-duration (124 to 199 days) ISS missions are presented. Immediately on return to 1 g, changes were observed in OPG and RANKL serum levels and the ability to produce unstimulated and stimulated PGA of peripheral blood mononuclear cells in vitro. Individual variability of these changes was noticed. Our findings suggest that the cytokine OPG/RANKL-system is involved in bone remodeling in members of long-duration space missions.

[Immune system in Russian cosmonauts after orbital space flights].

The article is an overview of the results of studies of the immune systems of cosmonauts. The use of a system approach to the evaluation of the various components of the immune system made it possible to identify a number of characteristics of adaptive change, including the quantitative and functional changes of the innate and adaptive immunity. Among them the most important are: changes in in the system of Toll like receptors (TLRs) manifested as a decrease in the content of circulating monocytes and granulocytes expressing TLR2, TLR4, and TLR6, and LPS-induced cytokine production; inhibition of the functional potential of natural killer (NK)- and T-cells. The article discusses possible factors and mechanisms of the identified changes.

[Immunological aspects of a piloted mission to Mars].

The paper deals with the results of the effects of 520-day isolation and confinement modeling some elements of a mission to Mars on the immune system. Longitudinal analyses revealed that the mechanisms of adaptive response of the human immune system to the conditions of extremely long isolation led to a change of the parameters, characterizing innate and adaptive immunity. Among them the most important are: changes in the signaling PRRs--TLR, manifested in the reduction of the percentage of circulating monocytes and granulocytes expressed on its own surfaces TLR2, TLR4 and TLR6, decreases early NK cell activation potential, increases in the percentage T- and B-lymphocytes, that expressed early activation marker CD69 after adequate stimulation, and in production of cytokines in response to PHA stimulation. The active mobilization of the mechanisms of adaptive immunity, the implementation of the function of the level of immunity to a qualitatively different level, apparently, should be taken as a sign of adaptive adjustment of an organism in response to the complex influence of unfavorable factors, aimed at the preservation of immune homeostasis.

Endothelial dysfunction markers and immune response indices in cosmonauts' blood after long-duration space flights.

Space flight factors are known to cause a malfunction in the human immune system and lead to damage to blood vessels. The hemostatic function of endothelium during space missions and its interaction with human immunity has not been determined so far. In this work, we investigated the markers of endothelial activation and damage (plasma concentrations of soluble thrombomodulin fraction (sTM), von Willebrand factor (vWF), highly sensitive C-reactive protein (hs-CRP)), as well as the level of D-dimer and compared them to the immunological parameters characterizing the state of human humoral and cellular immunity. The immune status of long-duration ISS crewmembers was assessed by whole-blood testing, and comprehensive postflight immune assessment included the analysis of leukocyte distribution. Flow cytometry was applied to determine the absolute counts and the percentage of lymphocyte subsets: B cells (CD19+), T cells (CD3+, CD3+CD4+, CD3+CD8+), NK cells (CD3-CD16+CD56+, CD11b+CD56+), and activated subsets (CD3+CD25+ and CD3+HLA-DR+). The in vitro basal cytokine production was investigated in whole blood cell culture. The cytokines IFN-gamma, IL-1-beta, IL-4, IL-6, IL-10, IL-18, and TNF-alpha were measured in plasma and the 24-h supernatants by a sensitive enzyme-linked immunosorbent assay. A significant increase in the plasma levels of vWF and hs-CRP and a decrease in the concentration of sTM after spaceflights were detected. Divergent changes in the parameters characterizing the state of the immune system were observed. We propose that the changes revealed may lead to an increase in the procoagulant activity of blood plasma, suppression of protein C activation and thrombin inhibition, as well as to an increase in the adhesive-aggregate potential of platelets, especially in case of changes in the rheological characteristics of blood flow during re-adaptation to ground conditions. We also speculate that the immune system might play an important role in vessel damage during long-duration missions.

The impact of short-term confinement on human innate immunity.

During space missions cosmonauts are exposed to a myriad of distinct stressors such as radiation, overloads, weightlessness, radiation, isolation in artificial environmental conditions, which causes changes in immune system. During space flights it is very difficult to determine the particular factor associated with the observed immunological responses. This makes ground-based experiments examining the effect of each space flight associated factor along of particular value. Determining mechanisms causing alterations in cosmonauts' immunity can lead to potential targets for different countermeasures. In the current article we present the study of the early period of adaptation of human innate immunity of 6 healthy test-subjects, 4 males and 2 females aged 25 through 40, to isolation factors (hypodynamia, psychological stress, artificial environment). We measured multiple parameters characterizing innate immunity status in blood samples at chosen time points before, during and after the mission. In the experiment, highly enhanced cytokine responses were observed upon ex vivo antigen stimulations in comparison to baseline values. For cellular parameters we found multidirectional dynamics with a persistent prevalence of increasing TLRs+ monocytes as well as TLRs expression. Our study provides evidence that even a short-term confinement leads to immune changes in healthy humans that may trigger aberrant immune response.

[Dynamics of blood concentration of neurospecific proteins and risk of neuropathy development in the conditions of 105-day confinement].

Six male volunteers (aged 25 to 40 years) were subjects in all-round psychophysiological, hormonal and immunological studies before, in and after 105-day isolation and confinement. Blood was drawn and the 16-factorial Cattell personality inventory was filled out every 30 days. Concentrations of blood hormones, neurospecific proteins and cytokines point to a close interrelation between antibody titers to myelin-associated glycoprotein and changes in the parameters of metabolism and reproduction-related hormones, as well as cytokines and individual psychophysiology (extra-introversion, dominance, intropunitiveness, social contact selectivity, etc.), and suggest a minimum risk of demyelinizing neuropathy due to exposure to the conditions of isolation and confinement.

[Congenital human immunity during 5-day dry immersion].

The system of congenital immunity was studied in 12 essentially healthy males 18 to 26 years of age subjected to 5-day dry immersion without use of countermeasures. Peripheral blood was analyzed for monocytes, granulocytes and lymphocytes expressing the TLR2+, TLR4+, TLR6+, CD11b+, CD14+, CD16+, CD18+, CD24+, CD36+, CD54+, CD56+ and CD206+ receptors. Expression of early activation marker CD69 on lymphocytes-natural killers was studied in unstimulated and interleukin-2 activated mononuclear cell cultures. The negative shifts in the congenital immunity system in some volunteers at the end of immersion and during recovery can be considered as warnings about depletion of the system reserve and increase of the risk of infectious diseases such as caused by normal microflora which typically does not provoke pathological reactions of the host.

[Analysis of the relationships between the psychophysiological status and system of adaptive immunity in the conditions of 5-day dry immersion].

Relationships of the T- and B-components of adaptive immunity and the psychophysiological status were studied in 14 volunteers for the experiment with 5-d dry immersion (DI) w/o countermeasures. Comparison of frequency of deviations in immunity parameters of psychologically different subjects demonstrated the highest frequency in non-anxious and extravert individuals on day-5 in DI. These differences in immune reactions as a function of psychological type and temperament point to existence of a neuroimmune typology and, therefore, the necessity of concurrent immunologic and psychological investigations in order to develop separate measures of rehabilitation from and prevention of stress in people with polar psychological status.

[Indicators of innate and adaptive immunity of cosmonauts after long-term space flight to international space station].

Results of innate and adaptive immunity indicators research at 12 cosmonauts who took part in long (128-215 days) expeditions to the International space station (ISS) are presented. It is shown that a space flight can lead to deflection of deviations in human immune system. These shifts occurred in decrease of phagocytes, NK, T-lymphocytes functional activity and also in abilities of immunocompetent cells to synthesize cytokines. Significant individual changes are noted in reaction of immune system to the long term space flight conditions specifying on individual predisposition to development of immune reactance infringements in the conditions of varying gravitational influences.

[Human immunity system status during 7-day dry immersion].

Subpopulations of lymphocytes, activation potential of T-, B- and NK-cells as well as cytokines production by immunocompetent cells in peripheral blood were studied in five volunteers for 7-day dry immersion without use of countermeasures. Results of the investigation revealed several negative shifts in the immunity and cytokines systems instigated by the experimental conditions. The considerable variability of the immune reactions to immersion suggests individual predisposition to immunological breaks in a changed gravity environment.

[Immune system parameters of healthy humans in a chamber experiment with hyperbaric oxygen-nitrogen-argon air].

The investigation into the immune effects of 9-d exposure to elevated pressures of normoxic and hypoxic O2-N-Ar atmosphere showed changes in a number of indices of genetic and adaptive immunity in normal humans. Dynamics and depth of the functional shifts under the influence of the experimental factors appear to be connected with genetically coded mechanisms of immunoreactivity. Monitoring of immune homeostasis is basic to the prevention of immune adaptation failure.

[Immunoglobulin e and allergen-specific IgE antibodies in cosmonauts before and after long-duration missions on the International Space Station].

Immunoglobulins (IgG, IgA, IgM, IgE), specific IgE-antibodies, and interleukin-4 (IL-4) were investigated in blood serum of nine cosmonauts before and after 128- to 195-day ISS missions. It was shown that long-duration space flight does not change significantly the content of serum immunoglobulins, allergen-specific IgE-antibodies or IL-4. Analysis for probable ratios of the total IgE and IL-4 contents in the pre- and post-flight periods did not reveal any linear correlation of these parameters.

Immune changes during long-duration missions.

Immune changes during space flights in excess of two weeks have been almost exclusively studied by Russian investigators. Most of these studies have compared postflight values with those obtained before flight. In a very few cases, analyses have also been attempted during flight or with samples collected during flight. Studies of cosmonauts during spaceflight have shown that IgG levels were unchanged, whereas IgA and IgM levels were sometimes increased. Additionally, inflight delayed type hypersensitivity testing demonstrated a decrease below the warning level in 1/3 of the cosmonauts tested. Pre- vs. postflight analyses have often revealed a postflight decrease in: PHA-triggered lymphocyte blast transformation; the proliferation index of T-lymphocytes in the xenogeneic graft versus host reaction; the mitogen-induced production of interleukin-2; the presence of certain leukocyte sub sets; and cytotoxic activity of natural killer cells. Other factors that either did not change, or changed in an apparently random manner after flight included: production of alpha and gamma interferon; autoimmune tests; and globulin classes.

Immune changes in test animals during spaceflight.

Over the past two decades, it has become apparent that changes in immune parameters occur in cosmonauts and astronauts after spaceflight. Therefore, interest has been generated in the use of animal surrogates to better understand the nature and extent of these changes, the mechanism of these changes, and to allow the possible development of countermeasures. Among the changes noted in animals after spaceflight are alterations in lymphocytic blastogenesis, cytokine function, natural killer cell activity, and colony-stimulating factors. The nature and significance of spaceflight-induced changes in immune responses will be the focus of this review.

Cell-to-cell interactions in changed gravity: ground-based and flight experiments.

Cell-to-cell interactions play an important role in all physiological processes and are mediated by humoral and mechanical factors. Mechanosensitive cells (e.g., osteocytes, chondrocytes, and fibroblasts) can be studied ex vivo to understand the effects of an altered gravity environment. In particular, cultured endothelial cells (EC) are very sensitive to a broad spectrum of mechanical and biochemical stimuli. Earlier, we demonstrated that clinorotation leads to cytoskeletal remodeling in cultured ECs. Long-term gravity vector changes also modulate the expression of surface adhesion molecules (ICAM-1, E-selectin, VCAM-1) on cultured ECs. To study the interactions of geterological cells, we cocultured endothelial monolayers and human lymphocytes, immune cells and myeloleucemic (K-560) cells. It was found that, although clinorotation did not alter the basal adhesion level of non-activated immune cells on endothelial monolayers, the adhesion of PMA-activated lymphocytes was increased. During flight experiments onboard the Russian segment of the International Space Station, we measured the cytotoxic activity of natural killer (NK) cells incubated with labeled target cells. It was found that immune cells in microgravity retained their ability to contact, recognize, and destroy oncogenic cells in vitro. Together, our data concerning the effects of simulated and real microgravity suggest that, despite changes in the cytoskeleton, cell motility, and expression of adhesion molecules, cell-cell interactions are not compromised, thus preserving the critical physiological functions of immune and endothelial cells.

[Modification of a method for determining the cytotoxic activity of lymphocytes--natural killers in space experiments].

A space modification of a method for determining the cytotoxic activity of natural killers (NK) uses cultures of human lymphocytes and re-inoculated suspension of tumorous myeloblasts K-562. The 3H-uridine labeled target-cells fixated in 0.1 % formalin solution proved to be best suited to the conditions prohibiting regular recharge of cultural medium and utilization of radioactive agents. The following practical conclusions were made from the space-flown experiment: pre-launch procedures (lymphocyte isolation and delivery to the launch site) and time period till dock must not be longer than 5 days (1) and during transportation to the orbiting vehicle cell cultures must be kept at 18-20 degrees C (2). The modified method can be employed to study various aspects of cell interaction such as the NK ability to destroy target-cells in suspension cultured exposed in microgravity.

Functional changes in neutrophils and psychoneuroendocrine responses during 105 days of confinement.

The innate immune system as one key element of immunity and a prerequisite for an adequate host defense is of emerging interest in space research to ensure crew health and thus mission success. In ground-based studies, spaceflight-associated specifics such as confinement caused altered immune functions paralleled by changes in stress hormone levels. In this study, six men were confined for 105 days to a space module of ~500 m(3) mimicking conditions of a long-term space mission. Psychic stress was surveyed by different questionnaires. Blood, saliva, and urine samples were taken before, during, and after confinement to determine quantitative and qualitative immune responses by analyzing enumerative assays and quantifying microbicide and phagocytic functions. Additionally, expression and shedding of L-selectin (CD62L) on granulocytes and different plasma cytokine levels were measured. Cortisol and catecholamine levels were analyzed in saliva and urine. Psychic stress or an activation of the psychoneuroendocrine system could not be testified. White blood cell counts were not significantly altered, but innate immune functions showed increased cytotoxic and reduced microbicide capabilities. Furthermore, a significantly enhanced shedding of CD62L might be a hint at increased migratory capabilities. However, this was observed in the absence of any acute inflammatory state, and no rise in plasma cytokine levels was detected. In summary, confinement for 105 days caused changes in innate immune functions. Whether these changes result from an alert immune state in preparation for further immune challenges or from a normal adaptive process during confinement remains to be clarified in future research.

Effect of SLS-2 spaceflight on immunologic parameters of rats.

During the Spacelab Life Sciences-2 mission, rats were dissected in space and biosamples were returned to Earth for analysis. Immunologic studies addressed the kinetics of T lymphocyte proliferative responses, cytotoxic activity of natural killer cells, and cytokine production. Experiments were performed by using spleen and bone marrow of rats dissected before flight, during flight, immediately after landing of the space shuttle (R + 0), or 14 days after landing (R + 14), as well as those of respective control animals. Each group consisted of five male Sprague-Dawley rats. It was demonstrated that T lymphocyte activity of rats dissected in flight was significantly decreased compared with the controls. This was observed during 48-, 72-, and 96-h cultivation and stimulation with the following mitogenic stimuli: concanavalin A (Con A; 0.1, 1.0, and 10.0 mg/ml), phytohemagglutinin (PHA; 2.5 mg/ml), and interleukin-2 (IL-2; 1 U/ml). The cell proliferation rate in rats dissected immediately after landing did not decrease, whereas that in rats dissected at R + 14 increased. The activity of spleen natural killer cells was reduced in response to 51Cr-labeled target cells during flight (YAC-1 and K-562) and after flight (YAC-1). At R + 14, their activity returned to normal. Another technique employed to measure natural cytotoxicity, using [3H]uridine-labeled target cells and ribonuclease, did not reveal any differences between control and experimental groups. In bone marrow, the activity of natural killer cells did not vary significantly. The production of IL-1, IL-2, tumor necrosis factor (TNF)-alpha, and TNF-beta in spleen cell cultures of the flight rats was reduced. At R + 0, IL-1 and TNF-beta levels remained lowered, whereas TNF-alpha was increased. At R + 0, interferon-alpha and interferon-gamma levels were diminished. In summary, cell-mediated immunity in rats was significantly suppressed during flight. The time course variation of immune parameters after flight suggests that the changes may truly indicate a response of the immune system to spaceflight conditions that could increase over time.