Young Sca-1+ bone marrow stem cell-derived exosomes preserve visual function via the miR-150-5p/MEKK3/JNK/c-Jun pathway to reduce M1 microglial polarization

Abstract

Background: Polarization of microglia, the resident retinal immune cells, plays important roles in mediating both injury and repair responses post-retinal ischemia-reperfusion (I/R) injury, which is one of the main pathological mechanisms behind ganglion cell apoptosis. Aging could perturb microglial balances, resulting in lowered post-I/R retinal repair. Young bone marrow (BM) stem cell antigen 1-positive (Sca-1⁺) cells have been demonstrated to have higher reparative capabilities post-I/R retinal injury when transplanted into old mice, where they were able to home and differentiate into retinal microglia.

Methods: Exosomes were enriched from young Sca-1⁺ or Sca-1^- cells, and injected into the vitreous humor of old mice post-retinal I/R. Bioinformatics analyses, including miRNA sequencing, was used to analyze exosome contents, which was confirmed by RT-qPCR. Western blot was then performed to examine expression levels of inflammatory factors and underlying signaling pathway proteins, while immunofluorescence staining was used to examine the extent of pro-inflammatory M1 microglial polarization. Fluoro-Gold labelling was then utilized to identify viable ganglion cells, while H&E staining was used to examine retinal morphology post-I/R and exosome treatment.

Results: Sca-1⁺ exosome-injected mice yielded better visual functional preservation and lowered inflammatory factors, compared to Sca-1^-, at days 1, 3, and 7 days post-I/R. miRNA sequencing found that Sca-1⁺ exosomes had higher miR-150-5p levels, compared to Sca-1^- exosomes, which was confirmed by RT-qPCR. Mechanistic analysis found that miR-150-5p from Sca-1⁺ exosomes repressed the mitogen-activated protein kinase kinase kinase 3 (MEKK3)/JNK/c-Jun axis, leading to IL-6 and TNF-α downregulation, and subsequently reduced microglial polarization, all of which contributes to reduced ganglion cell apoptosis and preservation of proper retinal morphology.

Conclusion: This study elucidates a potential new therapeutic approach for neuroprotection against I/R injury, via delivering miR-150-5p-enriched Sca-1⁺ exosomes, which targets the miR-150-5p/MEKK3/JNK/c-Jun axis, thereby serving as a cell-free remedy for treating retinal I/R injury and preserving visual functioning.

Keywords: Bone marrow Sca-1+ cell; Exosomes; Ischemia/reperfusion injury; MEKK3; miR-150-5p.