| Background and Objectives:Radiation-induced lung injury is one of the most common complications of thoracic radiation therapy,and its pathological manifestations are divided into two stages: the early stage occurs within 6 months of radiation therapy,that is,radiation pneumonitis(RP);the late stage occurs when radiation After 6 months of treatment,the radiation pulmonary fibrosis(Radiation pulmonary fibrosis,RPF).Once radiation-induced pulmonary fibrosis occurs,it is difficult to reverse,which seriously affects the quality of life of patients.None of the drugs currently used in clinical practice,such as hormones and antioxidants,can inhibit the development of RPF.Therefore,how to prevent or reduce the lung injury caused by radiotherapy is an urgent problem to be solved at present.Previous studies have found that when biological tissues receive low doses of ionizing radiation below0.5Gy,normal tissues can be induced to produce adaptive protective responses,and then become resistant to subsequent therapeutic doses of radiation,which may be the key to preventing or reducing radiation damage.Based on this,this study replicates a mouse model of radiation lung injury to observe the adaptive protective effect of low-dose pre-irradiation on radiation produced by therapeutic doses and the expression of factors related to radiation lung fibrosis,and to explore the possible mechanism of action.Methods:After 1 week of acclimatization,320 5-week-old female C57BL/6j mice were divided into control(0Gy),low-dose(0.5 Gy),high-dose(20 Gy)and low-dose pre-irradiation(0.5+20 Gy)groups by random numbering method,with 80 mice in each group.Mice in the 6-week-old low-dose group and low-dose preirradiation group were placed in the immobilization device under full consciousness for 0.5 Gy low-dose X-ray whole-body irradiation.2 weeks later,mice in the 8-week-old high-dose group and low-dose preirradiation group were anesthetized and exposed to 20 Gy high-dose X-ray chest irradiation,while control mice were exposed to mock irradiation(0 Gy).The weight of the mice and the number of mice that died unexpectedly were recorded daily after irradiation.All mice were treated according to the designed time points(24 hours,1 month,3 months and 5 months)after completion of the irradiation schedule,with 20mice/group treated at each time point.The lung tissue of mice was taken,and the left lung was fixed in 4%paraformaldehyde and then made into a wax block,which was stained with H&E and Masson’s trichrome,and its pathological dynamic changes were observed under a light microscope.The right lung was immediately frozen in liquid nitrogen and stored in-80℃ refrigerator,and the expression of fibrosis-related factors pro SP-C,HOPX,E-cadherin,vimentin,TGF-β1and β-catenin in lung tissues was analyzed by RT-q PCR and Western blot.Results:1.Changes in body weight and survival rate of mice: the weight of mice in 0Gy group and 0.5Gy group increased gradually with time,while the weight of mice in 20 Gy group increased only slightly from24 hours to 1 month after irradiation,and from 24 hours to 1 month after irradiation.From 1 month to 5months,it showed a gradually decreasing trend.The body weight of mice in the 0.5+20Gy group gradually increased from 24 hours to 3 months after irradiation,and reached a peak at 3 months after irradiation.In addition,at 1 month,3 months and 5 months after irradiation,the body weight of 20 Gy group was significantly lower than that of 0Gy group,0.5Gy group and 0.5+20Gy group,and the difference was statistically significant(P<0.01).In addition,the Log-Rank test was used to analyze the survival rate between the groups,and the mortality rate of mice in the 0.5+20Gy group was significantly lower than that in the 20 Gy group(P<0.01).2.The results of H&E and Masson trichrome staining showed that the single high-dose irradiation group(20Gy)showed varying degrees of intra-alveolar hemorrhage,hyaline membrane and epithelial cell hyperplasia,widening of alveolar septa,a significant increase in type II alveolar epithelial cells in the alveolar wall,and an accumulation of macrophages and collagen that phagocytosed particles,and with increasing irradiation time,the The degree of fibrosis became more pronounced with increasing irradiation time,while the histopathological changes of the lung in the 0.5+20Gy group were reduced at all time points after irradiation compared with the 20 Gy group.3.The m RNA and protein expression levels of pulmonary fibrosis-related factors: the m RNA and protein levels of pro SP-C and HOPX were higher in the 0.5+20Gy group than in the 20 Gy group,except for the pro SP-C protein in the 3-month and 5-month post-irradiation groups.In addition,the expression of the epithelial cell marker E-cadherin was significantly decreased in the high-dose 20 Gy group compared with the control group from 1 month to 5 months after irradiation,both at the m RNA level and at the protein level,while the expression was increased in the 0.5+20Gy group compared with the 20 Gy group.More significantly,the m RNA levels of TGF-β1 in the 20 Gy group were 5.8-13.6 times higher than those in the other groups at 5 months post-irradiation.At 5 months after irradiation,both m RNA and protein levels of β-catenin were much higher in the 20 Gy group than in the 0.5+20Gy group(P<0.01).Meanwhile,in the early post-irradiation period(24 hours and 1 month),vimentin protein levels were significantly higher in the0.5+20Gy group than in the 20 Gy group(P<0.05).Conclusion:Low dose pre-irradiation has the effect of reducing radiation lung injury,and its mechanism of action may be through regulating the expression levels of pro SP-C,HOPX,E-cadherin,vimentin,TGF-β1 and β-catenin and other fibrosis-related factors in mouse lung tissues,thus improving the tolerance of lung tissues to therapeutic doses of radiation,which provides a new therapeutic idea for clinical prevention of radiation lung injury. |
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