Small Extracellular Vesicles Derived from Altered Peptide Ligand‐Loaded Dendritic Cell Act as A Therapeutic Vaccine for Spinal Cord Injury Through Eliciting CD4+ T cell‐Mediated Neuroprotective Immunity

Abstract The balance among different CD4+ T cell subsets is crucial for repairing the injured spinal cord. Dendritic cell (DC)‐derived small extracellular vesicles (DsEVs) effectively activate T‐cell immunity. Altered peptide ligands (APLs), derived from myelin basic protein (MBP), have been shown to affect CD4+ T cell subsets and reduce neuroinflammation levels. However, the application of APLs is challenging because of their poor stability and associated side effects. Herein, it is demonstrate that DsEVs can act as carriers for APL MBP87‐99A91 (A91‐DsEVs) to induce the activation of 2 helper T (Th2) and regulatory T (Treg) cells for spinal cord injury (SCI) in mice. These stimulated CD4+ T cells can efficiently “home” to the lesion area and establish a beneficial microenvironment through inducing the activation of M2 macrophages/microglia, inhibiting the expression of inflammatory cytokines, and increasing the release of neurotrophic factors. The microenvironment mediated by A91‐DsEVs may enhance axon regrowth, protect neurons, and promote remyelination, which may support the recovery of motor function in the SCI model mice. In conclusion, using A91‐DsEVs as a therapeutic vaccine may help induce neuroprotective immunity in the treatment of SCI.


Figure S4 .
Figure S4.The isotype used for flow cytometry.The images showed isotype of IFN-γ, IL-4, IL-17A, and Foxp3 fluorescence antibody used for flow cytometry.

Figure S5 .
Figure S5.In vivo tracing of different DsEVs in SCI mice.The images showed the distribution of DiR-labeled different DsEVs in SCI mice after 6 hours of injection (n=3).

Figure S9 .
Figure S9.KEGG pathway enrichment analysis of spinal cord in the A91-DsEVs-treated group and the PBS-treated group at 7 and 14 dpi.(a) KEGG pathway enrichment analysis illustrated the downregulated signal pathways in the A91-DsEVs-treated group compared to the PBS-treated group at 7 dpi.(b) KEGG pathway enrichment analysis illustrated the downregulated signal pathways in the A91-DsEVs-treated group compared to the

Figure S10 .
Figure S10.Mice hind limb motor function was improved after A91-DsEVs treatment.The images showed a better recovery of hind limb function was achieved in mice from A91-DsEVs-treated group at 35 dpi.

Figure S11 .
Figure S11.The assessment of the biotoxicity of A91-DsEVs in vivo.(a) HE staining results showed the normal shape of major organs in different treated mice at 3 dpi.(b) The result showed the similar expression levels of LDH and ALT in mice liver from different groups.Data were expressed as mean ± SD. (* p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 as assessed by One-way ANOVA with Dunnett's multiple comparisons).