gms | German Medical Science

Objective: Failure of oligodendrocyte precursor cell (OPC) differentiation contributes significantly to failed myelin sheath regeneration (remyelination) in demyelinating diseases such as trauma, stroke and multiple sclerosis. Although the reasons for this failure are not completely understood, several lines of evidence point to factors present following demyelination that specifically inhibit differentiation of cells capable of generating remyelinating oligodendrocytes. We have previously demonstrated that myelin debris generated by demyelination inhibits remyelination by inhibiting OPC differentiation. The objective of the present study was to investigate intracellular signaling events that mediate the inhibition of neural stem-precursor cell differentiation into oligodendrocytes and to identify mechanisms by which the inhibition can be overcome. This is important for both transplant-mediated repair strategies as well as for regenerative treatments that target endogenous neural stem-precursor cells.

Methods: Primary rat neural stem-/precursor cells were plated on inhibitory myelin substrate and the involvement of Fyn-RhoA-ROCK assessed by immunoprecipitation, RhoA-Gtp assay and Western Blot. The activation of PKC was assessed by immunocytochemistry and confocal laser microscopy. Inhibition of the above mentioned cascades was mediated by small molecule inhibitors as well as by RNAi mediated gene silencing. For all assays at least three independent experiments were conducted. For statistical analysis standard ANOVA and t-tests were used.

Results: In the present study we show that the myelin-mediated inhibitory effects on OPC differentiation are regulated by statistically significant downregulation of Fyn-1 activation, and RhoA activation as well as by modulation of protein kinase C signaling. Furthermore, we demonstrate that inhibition of either ROCK-II or PKC is able to induce OPCs differentiation in the presence of myelin resulting in a significant increase in differentiating OPCs.

Conclusions: Our results provide a mechanistic link between myelin, a mediator of OPC differentiation inhibition associated with demyelinating pathologies and specific signaling pathways amenable to pharmacological manipulation, and are therefore of significant potential value for future strategies aimed at enhancing CNS remyelination.