12 December 2017

Safe dose

Small doses of X-ray radiation do not harm human stem cells

MIPT blog, Naked Science

Biophysicists have shown that after exposure to small (80 mGr) doses of X-ray radiation, stem cells remain viable, actively divide and do not accumulate DNA damage in the next generations.

Article Pustovalova et al. published in the journal Aging.

Sergey Leonov, Director of the MIPT Phys-tech School of Biological and Medical Physics, Head of the Laboratory for the Development of Innovative Medicines at MIPT: "The radiation dose of 80 mGr is the dose that a person often receives when imaging procedures of internal structures and processes of the body, such as computed tomography and X-rays, are often used in conjunction with cell therapy. Our research helps to make predictions of side effects and health risks in people undergoing increasingly used cell therapy at the same time as diagnostic radiation."

At the moment, the direction of regenerative medicine is developing rapidly and productively. Based on the use of stem cells, the technology is aimed at restoring and renewing damaged human tissues and organs. Stem cells have a high potential for reproduction, the ability to self-renew and differentiate, that is, to transform into different types of cells. Being in almost all organs and tissues of the adult body, they can recognize the site of damage, migrate into it, directly replace damaged cells and help healing. At the same time, it is believed that the active use in medicine of diagnostics based on ionizing radiation (computed tomography, mammography or X-ray) potentially contributes to the formation and accumulation of damage in stem cells and their subsequent transmission to cellular descendants. This entails the death of cells, their premature aging, as well as oncotransformation.


Figure 1: Micrograph of the nucleus of a human mesenchymal stem cell. From left to right: DNA of the cell nucleus stained with DAPI (blue); clusters (foci) of the yH2AX protein (red dots) marking DNA damage; superimposed microimages (merged).

Taking into account the lack of data on the effect of low doses of radiation on the manifestation of long-term effects in stem cells, an international group of scientists, including Andreyan Osipov from the Federal Medical Biophysical Center named after A. I. Burnazyan, Sergey Leonov and Anastasia Tsvetkova from the Moscow Institute of Physics and Technology, conducted a series of experiments. As a result, it was shown that exposure to low doses of radiation does not cause manifestations of genome instability, premature aging and accumulation of DNA damage in the offspring of irradiated cells.

The body's reaction to X-rays

With a normal X-ray examination, a person receives from 0.001 to 10 milligrees (mGr) radiation depending on the type of procedure. Doses up to 100 mGr are considered small, above 1000 mGr – large. The study of the effects of exposure to large doses of X-ray radiation has been done for a long time. It was found that they cause a dose-dependent increase in the amount of damage such as double DNA breaks, which then lead to cell death, malfunctions of genes responsible for suppressing tumor development and activation of oncogenes. However, until now, the question of the negative effects of small doses of X-rays that each of us receives during routine examinations is controversial. Currently, the world regulatory authorities have adopted the so-called "linear threshold-free model", which implies that an arbitrarily small dose of ionizing radiation is harmful to living cells. This is incorrect and does not correspond to reality, since we are all exposed to the natural radiation background, and its complete absence leads to a deterioration in the ability of cells to eliminate DNA damage.


Figure 2: A) arrows indicate dividing cells labeled with fluorescent dyes and having damage - double DNA breaks; B) the dependence of the number of dividing cells of the control group and cells irradiated with doses of 80 mGr and 1000 mGr for 11 generations.

Criteria for assessing the effects of small doses

The interest in the study of double DNA breaks is due to the fact that among the DNA damage caused by ionizing radiation, they are the most critical for the further fate of the cell. Repair or correction of these DNA damages occurs slowly, while double breaks that are not eliminated during this process lead to serious cytogenetic disorders, inactivation of tumor-suppressing genes or activation of oncogenes and cell death. For a long time there was no method for assessing the formation of double-stranded DNA breaks after exposure to low doses of radiation. Classical methods made it possible to see the effects of only large doses. Thanks to the development of immunocytochemistry, biophysicists have tools that allow not only to calculate the number of double DNA breaks formed after exposure to low doses of radiation, but also to recognize the mechanism of their distribution in the cell nucleus and recovery. Clusters of proteins involved in DNA correction, after "staining" with the help of antibodies labeled with fluorescent dyes, can be seen under a microscope in the form of brightly glowing dots, which are called foci. For example, one of these proteins that mark DNA damage is the modified histone protein uN2AH.

The fate of posterity

It is worth noting that there are two main ways to eliminate double gaps in the cell. One of them, homological recombination, is a slow but correct way that allows you to accurately restore lost information in the damaged DNA chain. Another way, a non-homologous connection of the ends, leads to the loss of genetic information and, as a result, the occurrence of errors and mutations. At the same time, 8 out of 10 ruptures formed in the irradiated cell are eliminated in a fast, but not exact way.

Scientists have found that stem cells 24 hours after irradiation at a dose of 80 mGr have a greater number of uN2AH foci than cells irradiated with a large dose of 1,000 mGr. However, such an increased content of uN2AX foci was observed only in dividing cells and was absent in resting cells (see Fig. 2). It is known that double DNA breaks can form normally during cell division. Such gaps are eliminated by the correct method of homological recombination. At the same time, if we follow the further fate of irradiated cells for 11 generations, it becomes obvious that the descendants of cells irradiated at a dose of 80 mGr do not differ from the descendants of non-irradiated cells. Moreover, in the offspring of cells exposed to a low dose of radiation, there were no manifestations of genome instability, changes in the processes of division and premature aging (see Fig.2 and Fig.3).


Figure 3: A) arrows indicate aging cells labeled with dyes (blue – cytoplasm, white – cell nuclei); B) the number of aged cells of the control group and cells irradiated with doses of 80 mGr and 1000 mGr for 11 generations.

Andreyan Osipov, Professor of the Russian Academy of Sciences, Head of the Department of Experimental Radiobiology and Radiation Medicine of the Burnazyan Federal Medical Center: "The conducted studies indicate that the presence of uN2AH foci in cultured human stem cells 24 hours after exposure to X-ray radiation at a dose of 80 mGr is associated with cell division processes and does not lead to long-term effects of radiation associated with aging. This is a very important conclusion, since the foci of uN2AX are currently actively used for biodosimetry of radiation effects. Misunderstanding of the biological significance of residual foci can lead to a significant overestimation of doses and the risk of exposure in small doses."

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