Selective cargo sorting in stem cell‐derived small extracellular vesicles: impact on therapeutic efficacy for intervertebral disc degeneration

Abstract Background Growing evidence has suggested the role of stem cell‐derived small extracellular vesicles (sEVs) in intervertebral disc degeneration (IVDD). The cargo sorting of sEVs, particularly miRNAs, may be influenced when the donor cell is subjected to oxidative stress. Here, we discovered that miRNAs containing specific motifs are selectively sorted into intraluminal vesicles within mesenchymal stem cells (MSCs) in response to oxidative stress. Methods Analysis of miRNA cargoes in sEVs derived from normal MSCs (C‐sEVs) or stressed MSCs (T‐sEVs) was conducted using miRNA sequencing. Differential expressed miRNAs in sEVs and the identification of motifs were evaluated through bioinformatics analysis. Protein binding was assessed using immunofluorescent staining and immunoprecipitation analysis. Additionally, RNA pull down and RNA immunoprecipitation (RIP) immunoprecipitation were employed to determine the binding between miRNAs and proteins. The effects of C‐sEVs and T‐sEVs on IVDD were compared by detecting the expression levels of phenotypic genes in vitro or histological evaluation in vivo. Results The sorting process of miRNAs is mediated by the nucleocytoplasmic transport of heterogeneous nuclear ribonucleoproteins, which in turn facilitates the phosphorylation of SNAP25 and promotes the transport and secretion of sEVs. Additionally, CHMP1B plays a role in membrane repair and protects against cell ferroptosis upon oxidative stress, concurrently affecting the release of sEVs. Notably, stem cell‐derived sEVs associated with ferroptosis impair the therapeutic efficacy for IVDD. However, the application of engineered sEVs containing a specific miRNA inhibitor exhibits the potential to reinstate the therapeutic efficacy for IVDD both in vitro and in vivo. Conclusions Taken together, our findings shed light on the mechanism of miRNAs sorting into sEVs and offer new insights for the optimization of sEV‐based treatments during intervertebral disc regeneration. regeneration.