Effect Of Interferon Regulatory Factor-1(IRF1)on The Progression Of Radiation Skin Injury And Its Mechanisms

Part I Effect of interferon regulatory factor-1(IRF1)on irradiated skin cellsObjectives:To study the effect of interferon regulatory factor-1(IRF1)on irradiated skin cells.Methods:Firstly,immunofluorescence and Western blot were used to detect the distribution of IRF1 in the cells and the expression of IRF1 in the cells after ionizing radiation.Western blot was used to detect the expression of IRF1 in skin cells after infection with Ad-IRF1 adenovirus,and the expression of IRF1 was detected by luciferase reporter gene system.CCK8 assay,clone formation assay and EDU cell proliferation assay were used.Effects of overexpression of IRF1 in skin cells on the proliferative capacity of skin cells;analysis of the effects of overexpression of IRP1 on apoptosis in cells by flow cytometry and lactate dehydrogenase cytotoxicity assay;The assay detects differences in ROS levels in skin cells that overexpress IRF1.CCK8 method was used to detect the effect of down-regulation of IRF1 in skin cells on skin cell proliferation ability;flow cytometry was used to detect the effect of IRF1 on apoptosis in skin cells.Results:In two different skin cell lines(fibroblast WS1,epithelial keratinocyte HaCaT),when the skin cells are subjected to ionizing radiation,the intracellular IRF1 will appear in the nucleus in a short time,and with the increase of radiation dose The expression level of IRF1 is increased,and the expression level of IRF1 increases with the time after ionizing radiation.After overexpression of IRF1 in skin cells,the proliferation ability of skin cells is inhibited,and the apoptosis rate is increased.When skin cells were subjected to ionizing radiation,the apoptotic rate of the skin cells overexpressing IRF1 was increased compared to the control group.After overexpression of IRF1 in skin cells,intracellular reactive oxygen species(ROS)levels are elevated.After down-regulating IRF1 in skin cells,there was no significant effect on the proliferative capacity of skin cells.Downregulation of IRF1 reduces the apoptotic rate of cells after skin cells are exposed to ionizing radiation.Conclusion:This study shows that ionizing radiation can induce an increase in the transcriptional ability of IRF1 in skin cells.Overexpression of IRF1 inhibits the proliferation and differentiation of skin cells,and increases the apoptotic rate of skin cells by inducing an increase in the level of reactive oxygen species in the skin cells.Down-regulation of IRF1 expression by rottlerin in irradiated skin cells can reduce the rate of apoptosis in cells.Therefore,IRF1 may be a key transcription factor involved in the progression of radiation-induced skin damage.Part II Effect of gene knockout on the progression of radiation-induced skin injuryObjective:To study the effect of interferon regu.latory factor-1(IRF1)gene knockout on the progression of radiation-induced skin injury.Methods:The model of radiation-induced skin damage in IRF1 knockout mice was established to observe the effect of IRF1 deletion on radiation-induced skin damage.HE staining was used to detect the effects of ionizing radiation on the skin of IRF1 knockout mice,heterozygous mice and wild type mice.Western blot was used to detect the expression of IRF1 in the skin of IRF1 knockout mice.Transmission electron microscopy was used to observe the changes of skin cell morphology in IRF1 knockout mice after exposure to ionizing radiation.Results:After exposure to ionizing radiation,IRF1 knockout mice had less radiation-induced skin damage than heterozygous mice and wild-type mice.The expression of IRF1 in the skin tissue of IRF1 knockout mice was lower than that of heterozygous and wild type mice;the results of HE staining showed that the skin of IRF1 knockout mice was subjected to ionizing radiation,and the subcutaneous appendages were compared.The heterozygous and wild-type mice were complete;transmission electron microscopy showed that the skin cells in the skin tissues of wild-type mice and heterozygous mice were damaged in the skin tissues of IRF1 knockout mice after ionizing radiation.serious.Conclusion:IRF1 deletion can effectively reduce radiation-induced skin damage and is expected to reduce radiation-induced skin damage by controlling the transcription of IRF1.Part ? Screening for RNA profiles and related pathways affected by IRF1 knockout through microarray analysisObjectives:To study RNA profile alteration in skin tissues affected by IRF1 exposed to ionizing radiation.Methods:RNA expression profiling by microarray analysis was used to screen for differentially expressed RNA in skin tissue of IRF1 knockout mice/wild-type mice after ionizing radiation.Results:In IRF1 knockout mice,after exposure to ionizing radiation,there were a total of 1145 differentially expressed RNAs in the skin tissue,specifically 295 differentially expressed mRNAs,of which 205 mRNAs were up-regulated and 95 mRNAs were down-regulated;A differentially expressed lncRNA,of which 715 IncRNAs were up-regulated and 69 lncRNAs were down-regulated;there were 140 differentially expressed CircRNAs,of which 134 CirRNAs were up-regulated and 6 CircRNAs were down-regulated.Differentially expressed RNA mainly affects NF-?B signaling pathway,P13K-AKT signaling pathway,TNF signaling pathway,prolactin signaling pathway and estrogen signaling pathway.Conclusion:The results of RNA expression profiling indicate that ionizing radiation can induce changes in multiple genes and signaling pathways after IRF1 deletion.