Figure 5. Transplantation of immature hiPSC-Astros into Rag1−/− mouse brains subjected to PVL.

(A) A schematic diagram showing the timeline for the in vivo experiments from postnatal day (P) 6 to 60.

(B) Left, a cresyl violet-stained coronal brain section at the level of the hippocampus, where analyses occurred. The red box outlines the white matter area used for immunohistochemical analyses of PVL insult. Right, a diagram showing the cell transplantation site, which is adjacent to the injured white matter area.

(C) Representative images showing that grafted hiPSC-Astros are identified by human nuclei (hN) staining at P11. The transplanted hiPSC-Astros are also identified by hCD44, but negative for MBP staining. The squared areas in the middle panels are enlarged in the bottom panels. Arrowheads indicate the hN⁺/GFAP⁺ cells. CC, corpus callosum; LV, lateral ventricle; and HIP, hippocampus.

(D) Upper panels, representative images showing that there are more Olig2⁺ oligodendroglial cells in the ipsilateral (Ipsi) side than in the contralateral (Contra) side brain in both vehicle and hiPSC-Astro groups. Transplantation of hiPSC-Astros further increases the expansion of Olig2⁺ cells in the Ipsi side. Lower panels, representative images showing that in vehicle group, MBP expression in the CC is only decreased substantially in the Ipsi side, with no significant changes in the Contra side. Transplantation of hiPSC-Astros promotes myelination in the Ipsi side and has no significant effect on the uninjured Contra side.

(E) The number of Olig2⁺ cells in the analyzed area (n = 4 for each group). Student’s t test, **P < 0.01, and ***P < 0.001, comparison between the Ipsi side versus the Contra side; and ^##P < 0.01.

(F) Fluorescence intensity (FI) of MBP staining (n = 4 for each group). Data represent the FI value normalized to the Contra side brain of vehicle group. Student’s t test, *P < 0.05 and ***P < 0.001, comparison between Ipsi side versus Contra side; and ^#P < 0.05.

(G) Representatives and quantification of Olig2⁺/CC1⁺ and Olig2⁺/CC1⁻ cells in the Ipsi side brains of vehicle and hiPSC-Astro groups. Arrows indicate Olig2⁺/CC1⁺ cells and arrowheads indicate Olig2⁺/CC1⁻ cells. Student’s t test, **P < 0.01, and ***P < 0.001, n = 4 for each group.

(H) Representatives and quantification showing that the majority of hN⁺ transplanted hiPSC-Astros are positive for human TIMP-1 (hTIMP-1) staining at P11. Scar bars, 50 μm. n = 4. Scale bars, 50 μm. Blue, DAPI-stained nuclei.