Background: Manipulation of neural stem and progenitor cells (NSPCs) is critical for the successful treatment of spinal cord injury (SCI) by NSPC transplantation, since their differentiation into neurons and oligodendrocytes can be inhibited by factors present in inflamed myelin. In this study, we examined the effects of LINGO-1 on spinal cord-derived NSPC (sp-NSPC) differentiation, the underlying mechanisms of action, and the functional recovery of mice after transplantation of manipulated cells.
Methods: sp-NSPCs were harvested from female adult C57/BL6 mice after SCI induced with an NYU impactor. These cells were infected with lentiviral vectors containing LINGO-1 shRNA sequence or a scrambled control and transplanted into SCI mice. Tuj-1- and GFAP-positive cells were assessed by immunofluorescence staining. Wnt5a, p-JNK, JNK, and β-catenin expression was determined by Western blot and RT-qPCR. miRNAs were sequenced to detect changes in miRNA expression. Motor function was evaluated 0-35 days post-surgery by means of the Basso Mouse Scale (BMS) and by the rotarod performance test.
Results: We discovered that LINGO-1 shRNA increased neuronal differentiation of sp-NSPCs while decreasing astrocyte differentiation. These effects were accompanied by elevated Wnt5a protein expression, but unexpectedly, no changes in Wnt5a mRNA levels. miRNA-sequence analysis demonstrated that miR-15b-3p was a downstream mediator of LINGO-1 which suppressed Wnt5a expression. Transplantation of LINGO-1 shRNA-treated sp-NSPCs into SCI mice promoted neural differentiation, wound compaction, and motor function recovery.
Conclusions: LINGO-1 shRNA promotes neural differentiation of sp-NSPCs and Wnt5a expression, probably by downregulating miR-15b-3p. Transplantation of LINGO-1 shRNA-treated NSPCs promotes recovery of motor function after SCI, highlighting its potential as a target for SCI treatment.
Keywords: Differentiation; LINGO-1; Neural stem and progenitor cells; Spinal cord injury; Wnt5a; miR-15b-3p.