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. 2021 Nov 5;38(1):99–103. doi: 10.1007/s12264-021-00783-9

Fig. 2.

Fig. 2.

The potential pathological role of AST487 in HD. A Chemical structure of AST487. B AST487 regulates HIPK3 kinase activity. The plots are fitted with a logistic curve. C Apoptosis in STHdhQ7/Q111 cells at different time points with or without AST487 treatment (active caspase 3 was measured using NucView 488 dye). Data are presented as the mean ± SD. D As in C, but using STHdhQ7/Q7 cells transfected with exon 1 HTT-Q72 plasmid. E Representative images and quantification of primary neurons (HDQ7/Q140), note that the mock group was the same for both the Fig. 1A and Fig. 2E quantification, scale bar, 10 µm. F Representative immunostaining results of the neuron-specific tubulin marker Tuj1 and DAPI showing the morphology of human iPSC-derived neurons (HD: Q47; WT: Q19) [note that the mock group was the same for both the Fig. 1B and Fig. 2F quantification; scale bar, 10 µm; data are presented as the mean ± SD (normalized to the WT)]. G Representative images and quantification of the caspase 3 signal in human iPSC-derived neurons (HD: polyQ = 47; WT: polyQ = 19), note that the mock group was the same for both the Fig. 1C and Fig. 2G quantification, scale bar, 10 µm. H mHTT levels in human iPSC-derived neurons (HD: polyQ = 47; WT: polyQ = 19). I mHTT levels in fly heads assessed by 2B7/3B5H10 using HTRF (fly model: fl-HTT-Q128). J Representative images and quantification of LC3B Western blots for Hipk3 RNAi or Hipk3 inhibitor AST487 treatment in mouse primary neurons (HDQ7/Q140) (n = 3 replicates). K As in J, but in human iPSC-derived neurons (HD: polyQ = 47) (n = 3 replicates). L Representative caspase 3 Western blots for Hipk3 RNAi or Hipk3 inhibitor in mouse primary neurons (HDQ7/Q140). M As in L, but in human iPSC-derived neurons (HD: polyQ = 47). Mock, N2 and B27 were not removed from the medium for mouse primary neuron culture, BDNF was not removed from the medium for iPSC-derived neuron culture. Data are presented as the mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; ns, P > 0.05; two-way ANOVA (C, D), one-way ANOVA and post hoc Dunnett’s tests (E–G), or the unpaired t-test (H–K).