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. 2017 Aug 7;8(49):85428–85441. doi: 10.18632/oncotarget.19976

Figure 4. U18 treatment leads to impairments of self-renewal and neuronal differentiation in WT-iNSCs through abnormal cholesterol accumulation.

Figure 4

(A) Representative filipin staining results indicates that U18 treatment induced cholesterol accumulation, compared to non-treated WT-iNSC, scale bar = 50 μm. U18 was treated at various concentration to determine the appropriate concentration. The intensity of filipin was analyzed and quantified. (B) U18-treated WT-iNSCs were stained with antibody against LAMP-1 and quantified using the same method as in Figure 3C, scale bar = 50 μm, zoom scale bar = 10 μm. (C) Phase-contrast images of the control- and U18-treated WT-neurospheres, scale bar = 100 μm (left). Self-renewal ability of U18-treated WT-neurosphere was characterized by the size (middle) and number (right) of neurospheres. (D) After U18 treatment, WT-iNSCs were differentiated into neurons (α-INTERNEXIN, NF, and TUJ1). Nuclei were counterstained with DAPI, scale bar = 50 μm, zoom scale bar = 10 μm. (E) U18-treated WT-iNSCs and non-treated WT-iNSCs were differentiated into GFAP-positive astrocytes. Nuclei were counterstained with DAPI, scale bar = 50 μm. Differentiation capacity into neurons (D) and astrocytes (E) was quantified using the same method as in Figure 2C. *P < 0.05, **P < 0.01, ***P < 0.005.