Fig. 6

The impact of small molecule inhibitors on the intracellular accumulation of Tau. The intracellular level of Tau in cells left untreated as control (UTC) or treated with 50 µM Chlorpromazine (CPZ), 20 µM Cytochalasin D (CD), 30 µM 5-N-ethyl-N-isopropyl amiloride (EIPA), 75 µM Dyngo-4a (DYNGO), 200 µM Genistein (GEN), or 10 µM Nystatin (NYST) for 30 min before incubation with A fluorescently labeled monomers (FL-Mono), and B fluorescently labeled small fibrils (FL-S-fib), both at 250nM concentration for 3 h (exceptionally, EIPA were present during the incubation with Tau). Error bars represent SEM; n = 9–14. One-way ANOVA ****p < 0.0001. Fluorescence measurement of cells treated with 25 nM fluorescently labeled Tau C FL-Mono, and D FL-S-fib in the presence of 100 nM bafilomycin A1 (Baf), 30 µM chloroquine (CQ), 100 nM MG132, 200 µM Atropine (Atr) or 20 µM Pirenzepine (Pirz) for 20 h. Error bars represent SEM; n = 9–14 independent experiments per experimental condition. One-way ANOVA ****p < 0.0001, *p < 0.05 vs. UTC. E Fluorescence analysis of iPSCNs treated with 25 nM labeled Tau Monomers and small fibrils for 20 h in the presence of 2 µM Heparin. Error bars represent SEM; n = 3. One-way ANOVA followed by post-hoc test; ****p < 0.0001 vs. UTC. ns: not significant. Kinetics of intracellular Tau accumulation in LUHMES neurons pre-treated with 100 µM Heparin for 2 h before exposure to F 250 nM fluorescently labeled Mono (FL-Mono) and G 150 nM fluorescently labeled-small fibrils (FL-S-fib). The significance was calculated between “No pretreat” and “Pretreat” at each time point (Only significant points were shown). Error bars represent SD. n = 3 per experimental condition. One-way ANOVA followed by posthoc test; *p < 0.05, **p < 0.01, ***p < 0.001 vs. “No pretreatment”. H Intracellular accumulation in LUHMES neurons pretreated with 100 µM Heparin for 2 h before 9 h treatment with 100 nM fluorescently labeled Tau Monomers (Mono), 50 nM Oligomers (Oligo), or 50 nM mixture of aggregates including large fibrils, small fibrils, and oligomers. Error bars represent SEM. n = 3 per experimental condition. One-way ANOVA followed by posthoc test; ***p < 0.001, ****p < 0.0001 vs. UTC. ns: not significant. I Representative images of H. Scale bar: 25 nm