Figure 5

IDS deficiency affects the number of PDGFR-α+ progenitors both in mouse and in human: potential therapeutic effects of PDGF. (a) Confocal microscopy images of wt and IDS-ko mice brain cortex immunostained for the glial PDGFRα marker. Scale bar: 75 μm. Inset images show the difference between the wt long-branching and the mutant round-shaped morphologies of PDGFRα+ cells. Scale bars: 26 μm in wt and 8 μm in IDS-ko. (b) Quantification of PDGFRα+ cells over total DAPI+ cells. (c) Confocal microscopy images of wt and Hunter human cortex immunostained for the glial PDGFRα marker. Scale bar: 75 μm. Inset images show the difference between the wt long-branching and the mutant disrupted morphologies of PDGFRα+ cells. Scale bars: 22 μm in healthy and 11 μm in Hunter. (d) Quantification of PDGFRα+ cells over total DAPI+ cells. (e and f) Graphs showing the proliferation rate of wt NSC (red, orange) and IDS-ko NSC (blue, light blue) when cultured in the presence of both EGF and FGF2 (E) or FGF2 alone (f) with or without addition of PDGF. (g) Enzymatic IDS activity assay in wt and IDS-ko murine NSCs and related supernatants after addition of IDS to the culture medium for 5 div, showing the rescue of IDS activity in IDS-ko cells. (h) Confocal microscopy images of wt and IDS-ko NSCs cultured and differentiated for 10 div with or without addition of IDS or PDGF to the culture media and immunostained with antibodies against PDGFRα and Lamp1. (i–k) Histograms showing the percentage of PDGFRα+ (i), Caspase 3+ (j) and ubiquitin+ (k) cells over total nuclei in untreated, PDGF- or IDS-treated wt and IDS-ko NSCs differentiated for 10 div.The differences among all the values were statistically not significant unless indicated (*P≤0.05, **P≤0.01, ***P≤0.001); Student’s t-test was applied for b, d and g, whereas one-way ANOVA followed by Student’s t-test for i, j and k experiments