Figure 4.

Cytoplasmic α-Syn Inclusions and Impaired Neuro-supportive Function in Mature OLGs Derived through Maturation-Induction of α-Syn PFF-Treated OPCs

(A) Time chart of the experimental procedure is displayed. Cells are incubated with α-syn PFFs (1 or 3 μM) for 24 hr followed by complete removal of extracellular α-syn PFFs and initiation of 7 days of maturation.

(B) Cytoplasmic α-syn inclusion is confirmed by confocal microscopy. The scale bar represents 20 μm.

(C) The cytoplasmic inclusions contain endogenous rat α-syn. The inclusions are also labeled with Thioflavin S staining. The scale bar represents 20 μm.

(D) Percentages of OLGs containing cytoplasmic inclusions labeled with both endogenous rat α-syn and Thioflavin S are compared between OLGs with and without α-syn PFF (1 μM) pretreatment before maturation. Mean ± SEM; n = 3, respectively; independent cultures; one-way ANOVA, ^∗∗p < 0.01.

(E) Immunoblot analysis reveals reduced myelin-associated proteins, MBP and TPPP/p25α, in OLGs pretreated with α-syn PFFs before maturation.

(F) Immunostaining of primary cortical neurons incubated with conditioned medium from OLGs reveals that reduced neuronal expressions of MAP2 and NeuN are induced by α-syn PFF (3 μM) pretreatment to OPCs before maturation. Each scale bar represents 50 μm.

(G–I) Viability of primary neurons is evaluated by the quantification of (G) WST assay, (H) MAP2-positive areas, and (I) numbers of NeuN-positive cells. Mean ± SEM; n = 4, respectively; independent cultures; one-way ANOVA, ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001.