Mechanism of Macrophage Extracellular Traps Involved in the Formation of Encapsulating Peritoneal Sclerosis

Background: Encapsulating peritoneal sclerosis (EPS) is one of the most severe complications of peritoneal dialysis and poses a serious threat to the lives of peritoneal dialysis patients. However, the mechanism of the development of EPS is still unclear, and there is a lack of effective treatment. Methods: We collected cells from the peritoneal dialysis effluent of EPS patients for single-cell transcriptome sequencing and used scanning electron microscopy and immunofluorescence to co-localize and identify macrophage extracellular traps(METs), while using fluorescent tracer cells to observe the "netting effect" of METs in vitro. We also used the EPS mouse model and PAD4 knockout mice for in vivo experiments. We stimulated fibroblasts with purified METs in vitro to explore the downstream molecular mechanism. Results: The ratio of myofibroblasts in the peritoneal cavity of EPS patients was significantly increased compared with that of other patients by using scRNA-seq analysis. In addition, the macrophage-fibroblast interaction was significantly increased and the proportion of pro-inflammatory macrophages (S100A8+ Macro) was significantly higher in EPS patients. Immunofluorescence also showed that fibroblasts and macrophages were the most predominant cellular components in the adhesion region of the abdominal. Intrestingly, the METs formation pathway was significantly enriched in this celltype. The level of METs was positively correlated with the severity of EPS. Both PAD4 knockdown and injection of DNase-I effectively reduced METs and adhesion in EPS mice. In vitro study showed that METs induced fibroblast activation via p38/MAPK pathway. Conclusion: Our study found that Macrophages in the peritoneal cavity produce METs under severe stimulation, and the reticular structure of METs can act as a scaffold to form "adhesion bridges" between cells in the peritoneal cavity and induce fibroblast activation through the p38/MAPK signaling pathway, which promotes peritoneal thickening and adhesion formation. Both knockdown of PAD4 and inhibition of METs by DNaseI were effective in improving EPS.