Fig. 2: Generation of position-specific brachial MNs corresponding to the MMC and LMC identity.

a Immunofluorescence analysis at day 20 in the soNMJ model revealed the presence of MYF5⁺ skeletal muscle progenitors and SOX1⁺ NPs. The presence of MNs was shown by the expression of ISL1 and ChAT. Scale bars: 75 µm. b Quantification of SOX1⁺ NPs (20.9% ± 7.6%), MNs expressing ISL1 (18.9% ± 10.1%), and skeletal muscle progenitors expressing MYF5 (58.4% ± 10.7%). ISL1 (H1: N = 3, n = 16; XM001: N = 3; n = 13); SOX1 (H1: N = 2, n = 6; XM001 N = 3; n = 9); MYF5 (H1: N = 2, n = 6; XM001: N = 2; n = 6) Source data are provided as a Source Data file. c Analysis of the MN population at day 20 revealed the presence of brachial MNs co-expressing HOXC6 and ISL1. Both LMC MNs expressing ISL1/FOXP1 and MMC MNs expressing ISL1/LHX3 were present in the soNMJ model. Scale bars: 50 µm. d Schematic illustration of MN columnar organization at the brachial spinal cord. e Quantification of HOXC6⁺/ISL1⁺ cells showing a prevalence of brachial MNs in the soNMJ model (80.2% ± 11.9%). Both MMC MNs (72.4% ± 14.4%), characterized by ISL1/LHX3 co-expression, and LMC MNs (25.3% ± 10%), characterized by ISL1/FOXP1 co-expression were present. ISL1 + HOXC6+ (H1: N = 3, n = 11; XM001: N = 3; n = 9; H9: N = 3, n = 9); ISL1 + LHX3+ (H1: N = 3, n = 9; XM001 N = 3; n = 9; H9: N = 1, n = 3); ISL1 + FOXP1+ (H1: N = 3, n = 9; XM001: N = 1; n = 3; H9: N = 2, n = 4). Source data are provided as a Source Data file. f Immunofluorescence analysis of brachial MNs expressing TUBB3, HOXC6, and ChAT at day 50. Scale bars: 50 µm. H1: N = 3, n = 9; XM001: N = 3, n = 9. g Schematic illustration of the neuromuscular connectivity in the human body at the forelimb level. soNMJ model: self-organizing neuromuscular junction model; NPs: neural progenitors; MNs: motor neurons; MMC: median motor column; LMC: lateral motor column.