a-d) 6-OHDA lesioned adult mice (NOD-SCID IL2Rgc null strain): a) TH expression and graft morphology at 4.5 months after transplantation. b) Expression of human specific marker (hNCAM, blue), TH (green), and FOXA2 (red). c) Quantification of FOXA2+ and TH+ cells in FP-derived grafts (mean ± SEM, n=4 at 4.5 months post grafting). d) Amphetamine-induced rotation analysis in FP- (blue) versus rosette-derived (green) grafts. Scale bars: 500 μm in (a), and 100 and 40 μm in (b).
e-p) 6-OHDA lesioned adult rats: Immunohistochemistry for TH (green) and human specific markers (red) hNA (e) and hNCAM (f). g) Stereological quantification of hNA+, TH+ and TH+ cells co-expressing FOXA2 (average graft volume = 2.6 ± 0.6 mm3). h-j) Co-expression of TH (green) with FOXA2, PITX3 and NURR1 (red). k-m) Behavioral analysis in FP- versus sham-grafted animals. k) Amphetamine-induced rotational asymmetry. l) stepping test: measuring forelimb akinesia in affected versus non-affected side. m) Cylinder test: measuring ipsi- versus contra-lateral paw preference. Grafted animals showed significant improvement in all three tests (p < 0.01 at 4.5–5 month; n=4–6 each). n-p) Immunohistochemistry for TH (green) and co-expression (red) with DAT (n), GIRK2 (o) and calbindin (p). Significance levels (panels d, k, l, m): ** p < 0.01; p < 0.05). Scale bars: 200 μm in (e), 50 μm in (f), 20 μm in (h-j) and 40 μm in (n-p).
q-t) Adult MPTP lesioned rhesus monkeys. q) Representative graft at 1 month after transplantation expressing human specific cytoplasm marker SC-121 (green). r) TH expression in graft with surrounding TH+ fibers (arrows). s) Co-expression of SC-121 (red) and TH (green). t) Co-expression of FOXA2 (red) and TH+ (green). Scale bars: 2mm for (q), 500 μm for (r), 200 μm for (s), and 50 μm for (t).