#4662 msc-derived exosomes prevent peritoneal fibroblast transdifferentiation in peritoneal dialysis-associated peritoneal fibrosis

Abstract Background and Aims Peritoneal dialysis (PD) as a long-term renal replacement therapy is limited by peritoneal fibrosis. The purpose of our study was to identify the mechanism of mesenchymal stem cell-derived exosomes (MSC-Exos) in regulating the transdifferentiation of fibroblasts (FBs) to myofibroblasts (MFBs) in peritoneal dialysis-associated peritoneal fibrosis. Method The experimental mice were divided into 3 groups according to treatment: control group (Saline), peritoneal injury group (2.5% glucose peritoneal dialysate + LPS), and exosome group (2.5% glucose peritoneal dialysate + LPS + MSC-Exos). After six weeks of modeling, the parietal peritoneum was collected for histological analyses and transcriptomic analysis. And we verified the effects of MSC-Exos on fibroblasts in vitro and in vivo experiments referred to immunofluorescence, quantitative real-time RT–PCR assay, and western blotting analysis. Results First, tracing study of DiI-labeled exosome showed that exosomes can enter peritoneal tissue and peritoneal cells in vivo and in vitro. Our studies have found that MSC-Exos can alleviate peritoneal fibrosis in peritoneal injury mouse model. MSC-Exos treatment significantly decreased the expression of mouse peritoneal MFBs markers (α-SMA), while maintaining the phenotype of fibroblasts (Collagen I), indicating that MSC-Exos might inhibit the transdifferentiation of FBs to MFBs. The in vitro results further confirmed these findings. So we supported that MSC-Exos could alleviate peritoneal fibrosis by inhibiting the transdifferentiation of FBs into MFBs. Transcriptomic analysis results revealed that SPIC gene which is related to cell differentiation ranked first in the differential expression genes. The expression of Spi-C was significantly increased in peritoneal injury group, while were significantly down-regulated in MSC-Exos group. The results showed that Spi-C overexpression or knockdown had no significant impact in MSC-Exos group, while the transdifferentiation of FBs into MFBs greatly increased with Spi-C overexpression and significantly decreased with Spi-C knockdown in peritoneal injury group. The results indicated that Spi-C might be a critical molecule to promote peritoneal FBs to differentiate into MFBs in peritoneal injury mice. Moreover, the miRNA sequencing results showed that miR-34b-3p is one of the main components of MSC-Exos, and it can specifically bind to SPIC gene sequence. The dual-luciferase reporter assay revealed that miR-34b-3p mimic significantly reduced the luciferase activity of Spi-C reporter. And miR-34b-3p mimics could down-regulate the expression of Spi-C, and the expression of Spi-C was not affected after miR-34b-3p inhibitor intervention. The above results suggested that MSC-Exos-derived miR-34b-3p could inhibit the transdifferentiation of FBs into MFBs by targeting Spi-C. Conclusion MSC-Exos-derived miR-34b-3p targeting Spi-C can alleviate peritoneal fibrosis by inhibiting the differentiation of subperitoneal mesothelial FBs into MFBs.