Fig. 6

Neurite outgrowth induction by 2D and 3D co-culture of hMSC and iMN in vitro. a Schematic of hMSC and iMN 2D and 3D co-culture neurite outgrowth assessment in vitro. b Representative light microscopy images of hMSC and iMN 2D co-culture into mature MN differentiation on day 10. c IF staining images of MAP-2 in 2D co-cultured hMSC and iMN on day 10. d Protein expression of MAP-2 in hMSC and iMN 2D co-culture cell lysates on day 10. Full-length WB images are presented in Additional file 7: Fig. S7. e IF staining of MAP-2 in 3D co-cultured spheroids consisting of hMSC and iMN on day 10. f Representative light microscopy images of neurite outgrowths on day 10 in the 3D co-cultured spheroids. g Representative fluorescence microscopy images of neurite outgrowths on day 10 in the 3D co-cultured spheroids. h Quantification of neurite outgrowth on day 10. Data are presented as mean ± SEM. Statistical significance was estimated using Kruskal–Wallis test with post hoc analysis and Mann–Whitney (†) test with least significant difference post hoc analysis (*); *, † P < 0.05. (WB analysis: hMSC n = 3, iMN n = 3, hMSC + iMN n = 3, Neurite outgrowth assay analysis: hMSC n = 6, iMN n = 6, hMSC + iMN n = 6, Scale bars = 20 μm. hMSC, human mesenchymal stem cells; iMN, induced pluripotent stem cell-derived motor neuron cells; IF, immunofluorescence; WB, Western blotting