Exosomes from WTAP-depleted mesenchymal stem cells mitigate OGD/R-triggered cellular injury in SK-N-SH cells through m6A-dependent epigenetic silencing of RPL9.

Background: Exosomes from mesenchymal stem cells (MSCs Exo) have emerged as a promising cell-free therapeutic strategy for human diseases, including ischemic stroke (IS). Here, we investigated the mechanisms underlying the therapeutic potential of MSCs Exo.

Methods: SK-N-SH cells were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R). Exosomes were isolated from untransfected (MSCs Exo) or shRNA-Wilms' tumor 1-associated protein (WTAP)-transfected MSCs (MSCs Exo-shWTAP) and used to incubate OGD/R-exposed SK-N-SH cells. The influence on cellular damage was evaluated by detecting cell viability, apoptosis, the expression of oxidative stress markers (ROS, MDA, and SOD), and the production of TNF-α, IL-6, and IL-1β cytokines. The regulation of WTAP in ribosomal protein L9 (RPL9) mRNA was assessed by mRNA stability analysis and methylated RNA immunoprecipitation (MeRIP) assay.

Results: RPL9 was upregulated in IS serum and OGD/R-exposed SK-N-SH cells. RPL9 depletion attenuated OGD/R-evoked apoptosis, oxidative stress, and pro-inflammatory cytokine production (TNF-α, IL-6, and IL-1β) in SK-N-SH cells. Mechanistically, WTAP regulated the mRNA stability and expression of RPL9 via an m6A-dependent way. MSCs Exo reduced RPL9 expression in OGD/R-exposed SK-N-SH cells, and MSCs Exo-shWTAP exerted a stronger reduction effect on RPL9 expression. MSCs Exo-shWTAP had stronger alleviative effects on OGD/R-triggered apoptosis, oxidative stress, and inflammation in SK-N-SH cells compared with MSCs Exo. Moreover, increased RPL9 expression abolished the effects of MSCs Exo-shWTAP.

Conclusions: Our findings indicate that WTAP depletion can enhance the alleviative effects of MSCs Exo on OGD/R-triggered cellular damage in SK-N-SH cells by downregulating RPL9. WTAP-depleted MSC-derived exosomes represent a promising therapeutic strategy for IS.