Exsomal microRNA-223 Attenuates Pyroptosis in Renal Ischemia/Reperfusion Injury by Targeting NLR Family Pyrin Domain Containing 3

Background: A recent study has reported that miR-223 inhibits renal ischemia/reperfusion (I/R) injury and pyroptosis. Mesenchymal stem cells derived exosomes (MSCs-exo) have been observed to improve renal functions after I/R injury, but the relationship between MSCs and miR-223 is not fully understood. Methods: Renal I/R mouse models were established. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-Ethynyl-2’-deoxyuridine (EdU) assays were used to assess cell viability and proliferation. Renal histopathologic changes were visualized using H&E staining. Enzyme linked immunosorbent assay (ELISA) was performed to assess interleukin (IL)-18, IL- 1β, serum creatinine, and blood urea nitrogen concentrations. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to assess RNA levels and western blot was used to assess protein levels. MSCs-exo characteristics were observed and evaluated using a transmission electron microscope (TEM) and nanoparticle tracking analysis (NTA). The targeting relationship was verified by luciferase reporter assay. Results: MSCs promoted mouse kidney tubular epithelium cells (TCMK-1) and miR-223 level viability, while the exosome inhibitor reversed this effect. In the I/R mice, pyroptosis was exacerbated, and miR-223 expression was inhibited. MSCs-exo exhibited circular/elliptical shape by TEM, with particle sizes mostly 85-110 nm. Exosome-specific proteins CD9 and CD63 surface antigens were positive in MSCs-exo, with higher levels than MSCs. It was verified that NLR Family Pyrin Domain Containing 3 (NLRP3) was the target gene of miR-223. Furthermore, NLRP3 specific inhibitor promoted cell viability, reduced pyroptosis related genes and cytokines in hypoxia/reoxygenation TCMK-1 cells, which were reversed by miR-223 inhibition. Additionally, NLRP3 - miR-223 interaction was confirmed in the I/R mice. Conclusions: Exosomal miR-223 participated in renal I/R-induced pyroptosis by targeting NLRP3, prompting a novel therapeutic strategy for kidney I/R injury.