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. 2018 Oct 19;27(1):87–101. doi: 10.1016/j.ymthe.2018.10.010

Figure 3.

Figure 3

Characterization of Rescue Compounds, Including Their Effects on c-jun Phosphorylation in Mouse Motor Neurons and Survival in Human Motor Neurons

(A) Dose-reponse curves for the lead compounds from the rescue screen (GÖ6976, n = 6 culture wells; K252a, n = 2 independent cultures; kenpaullone, n = 3 independent cultures; and TUDCA, n = 2 independent cultures). Bars denote average, and error bars indicate SEM. (B and C) Histogram (B) and confocal micrographs (C), showing the effects of rescue compounds on phospho-c-jun expression in CPA-treated motor neuron cultures at 2 hr of exposure. Bars denote average, and error bars indicate SEM; all groups were compared to CPA, ***p < 0.001 (one-way ANOVA, post hoc Dunnett’s multiple comparison test). Scale bar, 50 μm. (D) Proposed model for putative signaling pathways for CPA and targets for rescue compounds. Blue numbered labels indicate reported upstream or direct effects for the different rescue compounds. Intact lines show reported signaling pathways, and dashed lines show suggested pathways. Abbreviations are as follows: General control non-depressible 2, GCN2; Protein kinase R, PKR; and Heme-regulated inhibitor, HRI. Superscript numbers denote supporting references as follows: 1, Sakaki et al.67; 2, Lemonnier et al.85; 3, Kase et al.68; 4, Roux et al.69; 5, Sun et al.86; 6, Yang et al.13; 7, Meares et al.79; 8, Uppala et al.93; 9, Özcan et al.92; 10, Castro-Caldas et al.91; 11, present study. (E) Histogram showing the effects of CPA and in the absence or presence of rescue compounds (GO, GÖ6967; Kp, kenpaullone) in FACS-purified Hb9::GFP+ human motor neurons differentiated from an SOD1+/+ and an isogenic genetically modified SOD1+/A4V human ESC (hESC) line (n = 9 for CPA versus CTRL for both genotypes, and n = 3 for all rescue compounds for both genotypes). Bars denote average, and error bars indicate SEM; *p < 0.05, **p < 0.01, and ***p < 0.001 (unpaired two-tailed Student’s t test). (F) Representative cropped whole-well images showing calcein+ motor neurons. Scale bar, 50 μm.