During spaceflight, astronauts may be exposed to radiation that can compromise tissue function, even if the exposure is at a low dose rate. Multipotent precursor cells exist throughout the body and maintain the health of tissue by replenishing cells lost or damaged throughout life. Tissue can be compromised by age, disease or stressors, such as ionizing radiation. Studies have shown that relatively low doses of radiation found in space, such as protons and heavy ions, can compromise the function of specific tissues and organs through mechanisms involving oxidative stress.
Dr. Bertrand Tseng's project seeks to determine if and how relatively low doses of protons can initiate adaptive changes in precursor cells found in the brain and skeletal muscle system. The study will evaluate how low-dose exposure impacts precursor cell proliferation, differentiation and survival and whether such endpoints can be altered by prior exposure. One hypothesis is that neural or muscle precursor cell adaptation after a very low dose exposure enhances the cell's response to later exposures (e.g., from a solar flare). Another objective is to examine the relationship between this radioprotective adaptation and acute changes in oxidative stress. This relationship can be studied by using mice producing an enzyme that converts hydrogen peroxide into oxygen and water thus, decreasing oxidative stress.
Bertrand P. Tseng, M.D., Ph.D.
University of California, Irvine
Multipotent stem cells exist throughout the body and maintain the health of a given tissue by replenishing cells lost or damaged throughout life. These unique cells retain the capability to proliferate, migrate and differentiate to return functionality to a tissue compromised by age, disease or a variety of geno/cytotoxic stressors such as ionizing radiation.
Previously, we showed that relatively low doses of radiation found in space, such as protons and heavy ions, can compromise the function of specific tissues and organs through a variety of biochemical mechanisms that involve oxidative stress. Oxidative stress that persists after irradiation can reduce bone density, impair muscle re-growth and inhibit neurogenesis. These acute, functional decrements could severely compromise an astronautís ability to complete mission-critical tasks, where accelerated muscle fatigue and confusion caused by muscle and neural stem cell loss could result from relatively mild exposures to the space radiation environment. These debilitating conditions can, in part, be caused by radiation-induced oxidative stress. Our data suggest that these multipotent cells may represent critical targets for countermeasures aimed at ameliorating radiation and spaceflight-related sequelae.
Thus, we propose in vitro and in vivo experiments to determine if, and how, relatively low doses of protons can initiate radioadaptive changes in stem cells found in the brain and skeletal muscle. Our project will evaluate how low-dose proton exposure will impact stem cell proliferation, differentiation and survival and whether such endpoints can be altered by prior exposure. We hypothesize that stem cells incurring particle traversals during spaceflight can undergo adaptive changes that attenuate their response to subsequent larger exposures (e.g., from a solar flare). We will also investigate whether the mechanistic basis of adaptation involves changes in the redox environment such as oxidative stress. Specific Aims
To determine if/how low-dose proton irradiation elicits radioadaptive changes in neural precursor cells (NPCs) from the central nervous system that depend on oxidative stress.
To determine the acute dose response of satellite cells to proton irradiation with respect to survival, phenotypic fate and oxidative stress.
Elucidate how low-dose irradiation impacts neurogenesis in mice expressing human catalase targeted to the mitochondria (MCAT).
In Aims 1 and 2, a range of biochemical, oxidative and immunohistochemical approaches will be used that involve the quantification of reactive oxygen and nitrogen species, and the detection of oxidative damage in neural precursor and satellite cells. In Aim 3, we will utilize mice genetically modified to minimize mitochondrial hydrogen peroxide to determine how reducing intracellular oxidants impacts neurogenesis. These studies will help establish whether adaptation occurs after low-dose proton irradiation, and whether stem cells and oxidative stress represent logical targets for countermeasure strategies.
Key Findings To date, the key findings are that low-dose proton irradiation of rodent and human-derived NPCs and rodent satellite cells results in reactive oxygen species and nitrogen species levels that fluctuate over time, as well as a gradual increase in superoxide levels. Further, both dose and dose rate may be key components of radioadaption in the multipotent cells when examining survival/proliferation using our priming-challenge paradigm.
Plans for Coming Year The main impact of these findings is that we will focus on better understanding the radioadaptive effect of low-dose and low-dose rate proton irradiation on the NPCs and satellite cells. The plan for the coming year is to determine the dose limits and dose rate dependence of the priming dose with respect to eliciting radioadaptation, and the temporal interval during which the cells are radioadapted. Lastly, work will continue in determining whether or not the MCAT mice are radioprotected with respect to neurogenesis.
Bertrand P. Tseng, M.D., Ph.D.
University of California, Irvine
Very little is known about the low-dose and low-dose rate radiobiology of neural precursors cells. These studies will yield important information regarding how this radioresponse differs from the response to higher doses of ionizing radiation. Further, increased reactive oxygen species (ROS) can be damaging by challenging cellular repair and renewal processes, but it can also activate redox sensitive signaling pathways which may underlie an adaptive response in neural precursors. Our recent work has begun to elucidate the effectiveness of low-dose rate irradiation on the radioadaption of neural precursor cells, and it may prove to be quite beneficial as these dose rates better simulate the space radiation environment. Knowledge gained from understanding the mechanisms of change in oxidative stress, mitochondrial (MCAT) function and redox sensitive signaling in these experiments will provide the rationale for potentially effective countermeasure design for both acute and chronic space radiation exposure.
Ionizing radiation significantly reduces dentate neurogenesis, and such changes are dose dependent and persistent. These effects are linked to alterations in the neurogenic microenvironment, including inflammatory changes and oxidative stress although the precise mechanisms responsible are not yet known. While ROS have often been considered to be hostile or destructive entities, they also have been shown to have beneficial effects, at least in part due to their role as signaling moieties. It has recently been shown that a persistent level of oxidative stress in extracellular superoxide dismutase knockout mice (EC-SOD KO) was associated with a lower baseline level of neurogenesis relative to wildtype (WT) mice. However, when those same mice were subjected to a modest dose of x-rays (5 Gy), there was no effect on neurogenesis in KO mice but a significant reduction in WT mice. Thus, we saw both negative (baseline neurogenesis) and positive (adaptive) effects in the EC-SOD KO mice, presumably as a result of mechanisms associated with redox balance.
Also we have also been able to demonstrate the paradoxical nature of oxidative species in vitro using cultured neural precursor cells, where excess hydrogen peroxide reduces survival while excess superoxide increases survival. These paradoxical effects highlight the potential importance of adaptive responses in the context of the delicate balance in redox homeostasis, and how that may ultimately affect cell or tissue function. We believe that low doses of irradiation will elevate the level of oxidative stress in the neurogenic microenvironment and that this may have a beneficial effect when buffered by enhanced catalase activity with respect to dentate neurogenesis. These experiments can be conducted using a transgenic mouse with targeted expression of human catalase in the mitochondria (MCAT). Given the association between neurogenesis and cognitive function after irradiation, this modulation of oxidative stress via catalase could ultimately be developed into a countermeasure to maintain behavioral performance while engaged in space exploration.
With the capability to culture satellite cells in a multipotent state, we can now investigate the mechanistic details of ROS and reactive nitrogen species (RNS) production in live cells. Changes in ROS and RNS following proton irradiation may prove to be similar to what we have observed in either neural precursors exposed to the same irradiation at the same doses suggesting a common radioresponse pathway, or the changes might prove to be quite different, suggesting a more cell-type specific response. Similarly MCAT redox function via superoxide levels can be readily assayed in these. Understanding these effects of irradiation on myogenic oxidative stress, proliferation and differentiation will help to provide the basis for designing countermeasures to maintain muscle function during space exploration.