An estimate assay for low-level exposure to ionizing radiation based on mass spectrometry quantification of γ-H2AX in human peripheral blood lymphocytes.

Exposure to environmental ionizing radiation (IR) is ubiquitous, and large-dose exposure to IR is known to cause DNA damage and genotoxicity which is associated with an increased risk of cancer. Whether such detrimental effects are caused by exposure to low-dose IR is still debated. Therefore, rapid and early estimation of absorbed doses of IR in individuals, especially at low levels, using radiation response markers is a pivotal step for early triage during radiological incidents to provide adequate and timely clinical interventions. However, there is currently a crucial shortage of methods capable of determining the extent of low-dose IR exposure to human beings. The phosphorylation of histone H2AX on serine 139 (designated γ-H2AX), a classic biological dosimeter, can be used to evaluate the DNA damage response. We have developed an estimation assay for low-level exposure to IR based on the mass spectrometry quantification of γ-H2AX in blood. Human peripheral blood lymphocytes sensitive to low-dose IR, maintaining low temperature (4°C) and adding enzyme inhibitor are proven to be key steps, possibly insuring that a stable and marked γ-H2AX signal in blood cells exposed to low-dose IR could be detected. For the first time, DNA damage at low dose exposures to IR as low as 0.01 Gy were observed using the sensitive variation of γ-H2AX with high throughput mass spectrometry quantification in human peripheral blood, which is more accurate than the previously reported methods by virtue of isotope-dilution mass spectrometry, and can observe the time effect of DNA damage. These in vitro cellular dynamic monitoring experiments show that DNA damage occurred rapidly and then was repaired slowly over the passage of post-irradiation time even after exposure to very low IR doses. This assay was also used to assess different radiation exposures at the in vitro cellular level. These results demonstrate the potential utility of this assay in radiation biodosimetry and environmental risk assessment.

Differences in the Establishment of Gut Microbiota and Metabolome Characteristics Between Balb/c and C57BL/6J Mice After Proton Irradiation.

Although proton irradiation is ubiquitous in outer space as well as in the treatment of human diseases, its effects remain largely unclear. This work aimed to investigate and compare the composition of gut microbiota composition of mice in different species exposed to high-dose radiation. Male Balb/c mice and C57BL/6J mice were irradiated at a high dose (5Gy). Fecal specimens before and after irradiation were subjected to high-throughput sequencing (HTS) for the amplification of 16S rRNA gene sequences. We observed substantial changes in gut microbial composition among mice irradiated at high doses compared to non-irradiated controls. The changes included both the alpha and beta diversities. Furthermore, there were 11 distinct alterations in the irradiation group compared to the non-radiation control, including the families Muribaculaceae, Ruminococcaceae, Lactobacillus, Lachnospiraceae_NK4A136, Bacteroides, Alistipes, Clostridiales, Muribaculum, and Alloprevotella. Such alterations in the gut microbiome were accompanied by alterations in metabolite abundances, while at the metabolic level, 32 metabolites were likely to be potential biomarkers. Some alterations may have a positive effect on the repair of intestinal damage. Simultaneously, metabolites were predicted to involve multiple signal pathways, such as Urea Cycle, Ammonia Recycling, Alpha Linolenic Acid and Linoleic Acid Metabolism, Ketone Body Metabolism, Aspartate Metabolism, Phenylacetate Metabolism, Malate-Aspartate Shuttle, Arginine and Proline Metabolism and Carnitine Synthesis. Metabolites produced by proton irradiation in the microbial region play a positive role in repairing damage, making this area worthy of further experimental exploration. The present work offers an analytical and theoretical foundation to investigate how proton radiation affects the treatment of human diseases and identifies potential biomarkers to address the adverse effects of radiation. Importance: The space radiation environment is extremely complex, protons radiation is still the main component of space radiation and play an important role in space radiation. We proposed for the first time to compare the feces of Balb/c and C57BL/6J mice to study the changes of intestinal flora before and after proton irradiation. However, the effect of proton irradiation on the gut microbiome of both types of mice has not been previously demonstrated. After proton irradiation in two kinds of mice, we found that the characteristics of intestinal microbiome were related to the repair of intestinal injury, and some metabolites played a positive role in the repair of intestinal injury.

Combined Effects of Proton Radiation and Simulated Microgravity on the Cell Viability and ALP Activity of Murine Osteoblast Cells.

Proton radiation (PR) and microgravity (µG) are two key factors that impact living things in space. This study aimed to explore the combined effects of PR and simulated µG (SµG) on bone function. Mouse embryo osteoblast precursor cells (MC3T3-E1) were irradiated with proton beams and immediately treated with SµG for 2 days using a three-dimensional clinostat. All samples were subjected to cell viability, alkaline phosphatase (ALP) activity and transcriptome assays. The results showed that cell viability decreased with increasing doses of PR. The peak ALP activity after PR or SµG alone was lower than that obtained with the non-treatment control. No difference in cell viability or ALP activity was found between 1 Gy PR combined with SµG (PR-SµG) and PR alone. However, 4 Gy PR-SµG resulted in decreased cell viability and ALP activity compared with those obtained with PR alone. Furthermore, Gene Ontology analysis revealed the same trend. These results revealed that PR-SµG may lead to reductions in the proliferation and differentiation capacities of cells in a dose-dependent manner. Our data provide new insights into bone-related hazards caused by multiple factors, such as PR and µG, in the space environment.