Figure 6.

hPSC-derived reactive astrocytes also induce neurotoxic phenotypes upon cortical neurons and motor neurons

(A) Schematic demonstrating the derivation of both cortical neurons and motor neurons from hPSCs, with resulting cortical neurons identified by CTIP2 and MAP2 expression, while motor neurons could be identified by HB9 and βIII-tubulin expression. Images in Panel A created with BioRender.com.

(B–H) Cortical neurons were grown either in direct contact with astrocytes (B and C) or in transwell models (D and E). After two weeks of co-culture, reactive astrocytes resulted in a significantly decreased number of primary neurites from cortical neurons (F), a decrease in total neurite length (G), as well as in overall morphological complexity by Sholl analysis (H).

(I–O) Similarly, motor neurons were grown either in direct contact with astrocytes (I and J) or in transwell models (K and L). Morphological analyses demonstrated that reactive astrocytes promoted a reduction in the number of primary neurites in motor neurons, only in direct-contact culture system (M), as well as a significant reduction in total neurite length (N) and overall outgrowth complexity (O). Data represent mean values ±SEM from at least four independent experiments per hPSC line. One-Way ANOVA followed by Šídák’s multiple comparisons test with selected pair; ^∗p < 0.05, ^∗∗∗p < 0.001, ^∗∗∗∗p < 0.0001, and n.s. means non-significant, in (F), (G), (M), and (N). ^∗∗∗∗p < 0.0001, cortical neurons (in H) or motor neurons (in O) co-cultured with control astrocytes vs. reactive astrocytes in contact-dependent cultures, and ^####p < 0.0001, cortical neurons (in H) or motor neurons (in O) co-cultured with control astrocytes vs. reactive astrocytes in transwell cultures. Scale bar: 100 μm.