342 - Gut Microbiota and Radiation Induced Intestine Injury through Modulating Tissue Resident Macrophage Homeostasis

Z. Wang^1,2, J. Xiao^2,3, M. Cao^1,4, B. Xiang^1,2, D. Hu^1,2, X. Wang^1,2, Y. Wang¹, W. Liao¹, S. Zhang¹, J. Lang¹, and Y. Zhao¹; ¹Department of Radiation Oncology, Sichuan Cancer Hospital & Institute, Sichuan Cancer Center; Cancer Hospital affiliate to University of Electronic Science and Technology of China, Chengdu, China, ²School of Medicine, University of Electronic Science and Technology of China, Chengdu, China, ³Department of Radiation Oncology, Sichuan Cancer Hospital & Institute, Sichuan Cancer Center; Cancer Hospital affiliate to University of Electronic Science and Technology of China, Chengdu, Sichuan, China, ⁴Chengdu University of Traditional Chinese Medicine, Chengdu, China

Purpose/Objective(s): Radiation induced intestine injury significantly limits the application of radiotherapy in treating abdominal and pelvic malignancies. Intestinal macrophages constantly differentiate and polarize coincide with microbe metabolites, yet little is known how macrophage and microbiota orchestrate in shaping the outcome of radiation induced intestine injury. Intestinal resident CX3CR1+ macrophages are derived from embryos and replenished from peripheral circulating monocytes through a "monocyte waterfall" process depending on CCL2/CCR2 axis. The turnover kinetics and function of intestinal resident macrophage subpopulations governed by microbiota during radiation and the underlying mechanisms remains poorly defined. Herein, we used genetic modified mouse Ccr2^+/+ Cx3cr1^+/+ (WT), Ccr2^RFP/RFPCx3cr1^+/+ (R2-KO) and Ccr2^+/+Cx3cr1^GFP/GFP (X3-KO) mice in Abx treated murine radiation induced intestine injury model. Materials/

Methods: Mice were exposed to abdominal radiation of 13 Gy to induce radiation enteritis. Broad-spectrum antibiotics model (Abx model: ampicillin, neomycin, vancomycin and metronidazole) were used to investigate the functions of microbiota in regulating radiation enteritis. Severity of injury were evaluated and compared among groups through HE staining, immunohistochemistry, and electron microscope. Lamina propria immune cells were analyzed by flowcytometry. The gut microbiome from fresh fecal samples were detected by 16S rDNA sequencing, and the metabolites were detected by GC?MS analysis.
Results: Ionizing radiation increased the number of macrophages and co-expression of CCR2 and CX3CR1 in luminal propria, while does not alter the CD86/CD206 (M1/M2) ratio. Deletion of either CCR2 (R2-KO) or CX3CR1(X3-KO) significantly ameliorated radiation induced intestine injury as suggested by intestine length, length of villi, number of crypts and epithelial thickness. Abx pretreatment attenuated radiation induced injury in small intestine and colon, restored the length of villi and number of goblet cells. Recruitment of circulating monocytes to luminal propria macrophages pool after ionizing radiation relies on gut microbiota in that number of CCR2+ cells were significantly decreased in Abx treatment group. Notably, the turnover kinetics of CX3CR1^lo to CX3CR1^int macrophages and proportion of CD4⁺FoxP3⁺ was further upregulated in Abx treatment group. Mechanistically, Abx treatment enriched galactose-producing bacteria such as Ligilactobacillus, indicated by 16s rDNA. In addition, metabolomic profiling analysis suggested Abx-induced metabolic reprogramming, especially Biochanin A 7-sulfate enrichment in intestine.
Conclusion: Radiation induced intestinal resident macrophages turnover from CX3CR1^lo to dendritic-like CX3CR1^int macrophages is orchestrated by microbiota. Preferential supplement of microbiota and metabolites may provide novel insight to mitigate radiation induced intestine injury.