Novel Synthetic Coumarin-Chalcone Derivative (E)-3-(3-(4-(Dimethylamino)Phenyl)Acryloyl)-4-Hydroxy-2H-Chromen-2-One Activates CREB-Mediated Neuroprotection in Aβ and Tau Cell Models of Alzheimer’s Disease

Figure 3

Neuroprotective effects of coumarin-chalcone derivatives on Aβ-GFP-expressing SH-SY5Y cells. (a) Experimental flow chart. On day 1, cells were plated with retinoic acid (RA, 10 μM) being added to the culture medium. On day 2, the cells were treated with curcumin, LM-016, LM-021, or LM-022 (1.2–5 μM) for 8 h, followed by adding doxycycline (Dox, 5 μg/ml) to induce Aβ-GFP expression for 6 days. On day 8, Aβ-GFP fluorescence, ROS (CellROX Orange staining), neurite outgrowth (TUBB3 staining), and caspase 1/AChE activities were measured. (b) Analysis of GFP fluorescence with curcumin, LM-016, LM-021, or LM-022 (1.2–5 μM) treatment (; two-tailed Student’s -test; , , and ). The cell numbers counted for each treatment are shown in the following. For normalization, the GFP fluorescence/cell number of untreated cells (Untr) was set as 100%. (c) ROS assay with curcumin, LM-016, LM-021, or LM-022 (1.2–5 μM) treatment (). The cell numbers counted for each treatment are shown in the following. To normalize, the ROS/cell number of uninduced cells (Dox-) was set as 100%. (d) Aβ-GFP RNA, (e) caspase 1, and (f) AChE activity assays with curcumin, LM-016, LM-021, or LM-022 (5 μM) treatment (). To normalize, the caspase 1/AChE activities of uninduced cells (Dox-) were set as 100%. (g) Neurite outgrowth (length, process, and branch) assay with curcumin, LM-016, LM-021, or LM-022 (5 μM) treatment (). (c–g) values: induced (Dox+) vs. uninduced (Dox-) cells (^#, ^##, and ^###), or compound-treated vs. untreated (Dox+) cells (, , and ) (one-way ANOVA with a post hoc Tukey test).

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