scRNA-seq transcriptomic profiling of irradiated mouse skin reveals altered cell types,pathways,and cell-cell interactions

Objective:To investigate the substantial changes in cell types,pathways,and cell-cell interactions occurring in the irradiation-induced alopecia and dermatitis(IRIAD)mouse model and to identify potential targets for patients experiencing skin adverse reactions to radiotherapy.Methods:Mice were irradiated at 15 Gy,targeting the head and neck region.After a 14-day interval,living cells were extracted from both wild-type(WT)mice and irradiated mice for single-cell RNA sequencing(scRNA-seq).The scRNA-seq data,retrieved from the GEO database(GSE201447),underwent stringent quality control using the Seurat(v4.3.0)R package.Cell type annotation relied on previously reported typical markers and CellMarker 2.0.Differentially expressed genes were calculated to perform gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Cell-cell interactions were evaluated using the Cellchat R package.Results:The application of single-cell RNA sequencing(scRNA-seq)enabled a comprehensive characterization of the intricate cellular composition of both wild-type(WT)and irradiated mice skin.Remarkably,cells within irradiated mice skin exhibited a significant alteration in the intensity of cell-cell interactions compared to their wild-type counterparts.This change in interaction intensity was observed across various cell types,including fibroblast cells,endothelial cells,and dendritic cells.Importantly,these"interacting cells"shared common signaling pathways,notably the upregulation of the IL-17 pathway following irradiation.Conclusions:The modification of intercellular communication induced by irradiation primarily involves fibroblast cells,endothelial cells,and various types of immune cells.This investigation provides a novel perspective on potential targets and holds promise for enhancing the clinical management of IRIAD.

Effects of SWI/SNF complex on DNA damage repair in heterochromatin of embryonic fibroblast cells

Objective:To investigate the impact of SWI/SNF complex on heterochromatin DNA damage repair after exposure to X-ray irradiation,in order to explore the underlying mechanism.Methods:NIH3T3 and MRC5 cells were treated with 50 nmol/L siRNA targeting SWI/SNF complex subunits(BRM,ARID1A,BRG1 and SNF5),and YAP/TAZ.At 24 h after transfection,the cells were irradiated with 0.5 and 1 Gy of X-rays.At 20,60 and 240 min post-irradiation,γH2AX assay was performed to evaluate the radiation response in total or heterochromatin.Comet assay was used to determine the role of YAP/TAZ in DNA damage when the cells were irradiated with 4 Gy of X-rays.NIH3T3 were treated with 50 nmol/L siRNA targeting BRM/BRG1 and YAP/TAZ to determine their relationship on heterochromatin DNA damage repair.Results:In NIH3T3,SWI/SNF complex subunits(BRM,ARID1A and BRG1)knock-down increasedγH2AX in total and heterochromatin at 1 Gy 60 min post-irradiation(P<0.05),while SNF5 knock-down decreased heterochromatinγH2AX at 1 Gy 20 min post-irradiation(P<0.05).In MRC5,BRM and BRG1 knock-down increasedγH2AX in total and heterochromatin at 1 Gy 60 min post-irradiation(P<0.05).Inconsistently,ARID1A knockdown did not affect it,and SNF5 knock-down increased heterochromatinγH2AX at 1 Gy 60 min post-irradiation(P<0.05).Moreover,YAP/TAZ knock-down decreased heterochromatinγH2AX in NIH3T3 and MRC5(P<0.05).Meanwhile,YAP/TAZ knock-down decreased Tail Moment in comet assay at 4 Gy 60 min post-irradiation(P<0.05).BRM/BRG1 combining with YAP/TAZ knock-down significantly decreased heterochromatinγH2AX compared with single BRM/BRG1 knock-down at 0.5 Gy 60 min post-irradiation(P<0.05).Conclusions:The SWI/SNF complex subunits exhibited varying effects on DNA damage repair.BRM/BRG1 knockdown promotedγH2AX accumulation in heterochromatin through YAP/TAZ.This study provides a novel direction for DNA damage repair and sheds light on the role of SWI/SNF complex in response to DNA damage repair in heterochromatin.