Investigation of cellular senescence in the mouse liver caused by low dose fractionated X-ray therapy.

backgroundThe expanding use of medical diagnostics and radiotherapy has increased public exposure to low dose ionizing radiation (LDIR). A more clinically pressing concern, however, is the unintended irradiation of normal tissues adjacent to the radiotherapy target. Notably, the biological effects of lower cumulative doses on the liver remain poorly characterized.

methodsThirty-two 6 week-old C57BL6/J mice, half male and half female, were selected and randomly divided into control, 0.1 Gy, 0.2 Gy, and 0.5 Gy groups according to body weight and radiated every 48 hours for a total of 16 times. The body weights of mice were recorded 24 h after the end of each radiation exposure. The liver histopathology and liver index of mice were examined. The number of peripheral blood cells in mice was counted and serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), ceruloplasmin (CP) and C-reactive protein (CRP) levels were determined. Mouse liver tissues were screened for differentially expressed genes using high-throughput sequencing and subjected to gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and gene set enrichment analysis (GSEA) analyses. The senescence - associated SA-β-Gal staining as well as mRNA levels of cellular senescence genes, anti - apoptotic factors, and senescence associated secretory phenotypes (SASPs) were detected. The levels of oxidative stress and oxidative damage related indexes such as superoxide dismutase (SOD), malondialdehyde (MDA), reduced glutathione (GSH), and 8-hydroxy-2’-deoxyguanosine (8-OHdG) were detected and correlated with the levels of inflammation, oxidative damage level, and cellular senescence indicators level in mouse liver.

resultsCompared with the control group, the difference in body weight of mice in different dose groups did not show significant changes (p > 0.05). Compared with the control group, hepatocytes in the 0.1 Gy × 16 group showed mild edema; hepatocytes in the 0.2 Gy × 16, and 0.5 Gy × 16 groups all showed mild to moderate edema, with structural disorganization of the hepatic lobules, hepatic cords, central veins, and mild steatosis. The results of liver indices showed no significant changes in the 0.1 Gy × 16 and 0.2 Gy × 16 groups (p > 0.05) and a significant decrease in the 0.5 Gy × 16 group (p < 0.05) compared to the control group. As the radiation dose increased, leukocyte, neutrophil, lymphocyte counts, C-reactive protein and CP levels all decreased significantly (p < 0.05), while serum AST levels increased significantly (p < 0.05). Transcriptomic analysis of mouse liver showed that there were 46 genes differentially expressed across all dose groups. GO analysis showed that the differentially expressed genes in each dose group were mainly enriched in biological processes such as reactive oxygen metabolism process, NF-κB signaling pathway, and exogenous stimulus response. KEGG analysis showed that the pathways differentially expressed in the liver of mice in each dose group were mostly significantly enriched in the p53 signaling pathway, TNF signaling pathway, and IL-17 signaling pathway. GSEA-GO analysis showed that the 0.1 Gy × 16 group was enriched in biological processes such as acute inflammatory response, and the 0.2 Gy × 16 and 0.5 Gy × 16 groups were enriched in cell cycle regulation. The degree of SA-β-Gal staining of liver was increased in all dose groups compared to the control group. Compared with the control group, p53, p21, p16 mRNA and protein levels were significantly higher in the 0.2 Gy × 16 and 0.5 Gy × 16 groups (p < 0.05); IL-8, GM-CSF, TGF-β1, CXCL-1, CXCL-12, MMP-14, and MMP-15 mRNA levels were significantly elevated (p < 0.05). Compared with the control group, the level of the anti-apoptotic factor BCL2L1 mRNA was significantly increased in all dose groups (p < 0.05), the levels of MDA, SOD and GSH in the liver and the levels of 8-OHdG in both serum and liver of mice increased with increasing radiation dose. The levels of systemic inflammation and oxidative damage in mice were significantly correlated with the levels of indicators related to cellular senescence in liver (p < 0.05).

conclusionsLow dose fractionated X-ray therapy induces cellular senescence in mouse liver tissues, and the levels of systemic inflammation and oxidative damage in mice are correlated with indicators related to cellular senescence in liver tissues.