Regulation of CKD-504 on acetylated α-tubulin, mitochondrial transport, and mutant HTT accumulation using YAC128 NSCs. (A) Immunocytochemistry of acetylated α-tubulin. The WT and YAC128 NSCs were treated with CKD-504 (0.3, 1 μM) and vehicle (Veh), and CKD-504 (0.1, 0.3, 1, 3 μM), vehicle (Veh) and TBA (0.3 μM), respectively, for 12 hr. Scale bars: 50 μm. (B, C) Western blot analysis of acetylated α-tubulin in YAC128 NSCs treated as above (B) and its quantification (C, n = 3). (D, E) Representative kymographs of mitochondrial movement (D) and calculated mitochondrial velocity (E) in each group. Cells were treated with CKD-504 (0.1 μM) and vehicle for WT and CKD-504 (0.1, 0.3, 1 μM), vehicle and TBA (0.3 μM) for TG for 3 hr (n = 4). (F) ICC showing the decrease of mHTT aggregates that was detected by EM48 antibody after treatment as indicated for 12 hr (Scale bars: 20 μm). (G) Western blot analysis showing the decrease of EM48 positive HTT aggregates by the treatment of CKD-504 and TBA (n = 3). Data are presented as mean ± SEM. *P < 0.05: **P < 0.01: ***P < 0.001.
