Figure 2.

Representative photomicrographs of hiPSdNP transplanted into the cerebellum of adult NOD-SCID mice, time epochs are organized in columns. (a, d and g) Cerebellum sagittal sections. At increasing survival times the portion of cerebellum containing eGFP positive cells derived from the transplant increases due to cell dispersal and proliferation of neural precursors and glial cells. Scale bars: 100 µm. (b) Mouse cerebellum, at short survival times (15 days after the transplant) the majority of the engrafted cells maintain an undifferentiated morphology. Some cells bear distinct processes. Scale bar: 20 µm. (c) Mouse cerebellum survival as in B, sagittal section stained with anti hNestin (upper image) and anti NeuN (lower images) antibodies. Arrows mark examples of neurites double-labeled by antibody against eGFP and hNestin, asterisks indicate corresponding positions in all pictures, eGFP positive hiPSdNP derived cells are NeuN negative, while many of the surrounding host cells are NeuN immunopositive. Scale bars: 20 µm (upper), 10 µm (lower). (e) Mouse cerebellum 3 months after the transplant neuronal differentiation is ongoing with many process bearing cells immunopositive for hNCAM. The boxed area is enlarged at the bottom in order to show the close correspondence of eGFP and hNCAM positivity in neurites. Arrows indicate processes double positive for eGFP and hNCAM. Scale bars: 10 µm (upper), 5 µm (lower). (f) Mouse cerebellum 3 months after the transplant many neuronal cells are immunopositive for ß-III tubulin (arrows). Scale bar: 15 µm. (g) Mouse cerebellum 288 days after the transplant, higher magnification pictures of the areas labeled by the letters are provided below the low magnification image, showing the different morphologies and migration pattern of the cells derived from the transplanted hiPSdNP. Scale bars: 100 µm (upper image), 30 µm (lower four images). (h) Same survival as in G, many transplant derived cells differentiated into SOX10 positive cells (arrows). SOX10 is considered a marker of immature oligodendrocytes and myelinating oligodendrocytes⁵³ Scale bar: 15 µm. (i) Same survival as in G, examples of eGFP positive hiPSdNP derived cells immunopositive for GFAP showing an astrocytic morphology. Scale bars: 10 µm. (j) Same survival as in G eGFP positive hiPSdNP derived neurons (arrows) positive for hNCAM with extensive processes (arrowheads) are also present at these survival times. Scale bar: 10 µm.