Fig. 6.

Human iPSC-derived motor neurons produce neuroprotective factors. (A to D) Amounts of (A) NT3, (B) NT4, (C) NGF, and (D) VEGF secreted in vitro by motor neurons and evaluated by ELISA. **P < 0.00001, two-tailed Student’s t test. Mean ± SD, n = 12 independent experiments for each cytokine. (E and F) Average length of axons (E) and growth cone area of motor neurons (F) from SMA mice, in coculture with human motor neurons. Motor neurons from SMA mice cocultured with human motor neurons exhibited an increase in axon length and size of growth cone with respect to SMA mouse motor neurons not cocultured with human motor neurons (**P < 0.00001, two-tailed Student’s t test; mean ± SD, n = 4 independent experiments for each condition). (G) The average axonal length of cocultured SMA mouse motor neurons was reduced after cytokine neutralization of cytokine production by human motor neurons (P < 0.05, two-tailed Student’s t test; mean ± SD, n = 4 independent experiments for each condition). (H and I) Quantification of the number of SMI32-positive SMA murine motor neurons in the presence of human motor neurons and microglial-conditioned media with (I) and without (H) lipopolysaccharide (LPS). The number of SMA mouse motor neurons in cultures with microglia + LPS was reduced but was increased in coculture with human motor neurons (mean ± SD, n = 4 independent experiments for each condition). (J and K) Representative SMI32 staining of SMA murine motor neurons (black and white were used to show the morphometric characteristics, that is, number and axonal length of cells) exposed to microglial-conditioned medium with LPS, with (J) or without (K) human motor neuron coculture. Scale bar, 50 μm.