br Results br Discussion The transplant created

Results
Discussion The transplant-created chimera enables us to investigate the effects of human neural purchase Wortmannin in the adult mouse spinal cord (Chen et al., 2015). Using this model, we have shown that neural cells from sALS patient iPSCs, especially astrocytes, integrate into the mouse spinal cord to a similar degree as healthy cells. It is the sALS cells that induce degenerative changes in both MNs and non-MNs of the host, which corresponds to the mouse motor behavioral deficits. By taking advantage of the time-dependent integration of human cells in this model, we have discovered that non-MNs are lost earlier than MNs, with a corresponding reduction of inhibitory nerve terminals in MNs. Thus, the effect of sALS astrocytes on neural degeneration is not specific to MNs, and non-MNs may mediate MN degeneration. Involvement of glial cells in the pathogenesis of ALS has been suggested from a series of studies where disease-causing proteins are specifically expressed in glial cells in transgenic animals (Yamanaka et al., 2008) and when MNs are cultured with glial cells that express mutant ALS proteins (Di Giorgio et al., 2007, 2008; Nagai et al., 2007). The involvement of glia in sALS is suggested by recent observations that astrocytes, derived from neural progenitor cells that were reprogrammed from sALS patient fibroblasts (i-astrocytes), impaired the survival of MNs (Meyer et al., 2014). Nevertheless, astrocytes generated in a similar manner (via iPSCs) had no obvious effects (Re et al., 2014; Serio et al., 2013), casting a possibility that the in vitro toxic effects of astrocytes may be influenced by culture conditions. Our present study offers in vivo evidence that non-MN cells, especially astrocytes, may participate in the neural degeneration in sALS. This is demonstrated by the fact that neural progenitors differentiated from sALS patient but not healthy PSCs, generate non-MN cells following transplantation into the spinal cord of SCID mice, and cause neuronal degeneration and corresponding motor deficits in mice. It should be noted that one of the control transplants was performed at a different time yet the result was very similar, highlighting the consistency of the effect of the astrocytes and the reproducibility of the experiments. Furthermore, we show here that cells from both sALS patients have similar effects. ALS is heterogeneous. Although we excluded the possibility of C9orf72 mutations, the main contributor of genetic components to sALS, we cannot rule out other albeit extremely rare mutations and those that have not yet been discovered. Since the majority of the differentiated cells are astrocytes, astrocytes may play important roles in neuronal degeneration. In addition, the neuronal degeneration occurs mostly in the transplant center or near the center but not the distal area. Astrocytes in and around the transplant center are more mature than those in the distal area. This suggests that mature but not immature astrocytes exert toxic effects. This may in part explain why astrocytes generated from ALS patient iPSCs have limited effects (Re et al., 2014; Serio et al., 2013). Astrocytes generated from human ESCs or iPSCs usually exhibit immature phenotypes (Krencik et al., 2011). Co-culture studies have suggested a specific effect on MNs by ALS astrocytes (Di Giorgio et al., 2007, 2008; Nagai et al., 2007). The reason for the specific effect of ALS astrocytes on MNs remains a mystery. One possibility is that MNs are particularly vulnerable to toxicity, especially in the cell culture environment. In some cases, such a phenomenon may be misinterpreted due to the fact that changes in MN but not non-MN numbers are readily discerned because MNs are usually the minority in the culture system. Because of this mystery, we have paid a particular attention to the effect on MNs and non-MNs. Contrary to the in vitro observations, we found that both MNs and non-MNs degenerate in the spinal cord transplanted with sALS cells. This result suggests that the effect of astrocytes is not specific to MNs. An attempt to identify MN-specific astrocytic factors may be elusive.