Fig. 5.

Transplanted motor neurons ameliorate the disease phenotype of SMA mice. (A) Engraftment of human donor iPSC-derived motor neurons in spinal cord of SMA mice. Representative images of GFP-tagged transplanted motor neurons, located in the anterior horns of the spinal cord, coexpressed motor neuron– specific proteins like ChAT. GFP (green), ChAT (red), merged (yellow); nuclei were stained with DAPI (blue). Scale bars, 75 μm (first to second rows); 50 μm (third row). The engraftment analysis was performed at the disease end stage; n = 24 per group. (B) Quantification of engrafted motor neurons and donor axons in the anterior roots; n = 24 per group. (C) Representative image of neuromuscular junctions formed with paravertebral muscles by axons (green) of donor transplanted treated SMA-iPSC–derived motor neurons. Cholinergic receptors are labeled with bungarotoxin (BTX, red). Merge, yellow. Scale bar, 20 μm. (D) Human motor neuron transplantation ameliorated muscle atrophy in SMA mice. Histograms of myofiber diameters: Mean tibialis anterior muscle cross-sectional area was reduced in vehicle-treated SMA mice compared with WT littermates (P < 0.00001) and was augmented after transplantation of motor neurons from different sources (P < 0.00001). Data represent mean values ± SD. Mean total myofiber number was reduced in vehicle-treated SMA mice compared with WT littermates (P < 0.00001) and increased after transplantation of motor neurons from different sources (P< 0.00001). n = 6 per group at postnatal day 13 (P13). (E) Gross appearance of an SMA mouse transplanted with motor neurons derived from corrected SMA-iPSCs (MNTR), a vehicle-treated SMA mouse (SMA), and a WT mouse showing that the transplanted SMA mice were larger than vehicle-treated SMA mice (P13). Survival was extended for mice transplanted with motor neurons compared with vehicle-treated and fibroblast-transplanted SMA mice (P < 0.00001) as shown by Kaplan-Meier survival curves. Transplanted SMA mice survived for HET-MN (median, 21 days), SMA-MN (19 days), oligodeoxynucleotide-corrected SMA-MN (TR-MN, 21 days), or human primary fibroblasts (13 days) or vehicle-treated mice (SMA, 14 days). Transplanted SMA mice presented increased weight compared with vehicle-treated SMA mice (P13; P < 0.00001), as shown by weight curves. All plots are shown as means of weight with error bars representing SD. The grip time was statistically different between the transplanted and vehicle-treated SMA mice (P < 0.00001) at 13 days. Error bars represent SD. At P13, the number of crossings of transplanted mice was increased with respect to vehicle-treated mice in the open-field test (P < 0.00001, ANOVA followed by a Tukey’s post hoc test for multiple comparisons).