Oxidative Medicine and Cellular Longevity / 2020 / Article / Fig 6

Research Article

New Synthetic 3-Benzoyl-5-Hydroxy-2H-Chromen-2-One (LM-031) Inhibits Polyglutamine Aggregation and Promotes Neurite Outgrowth through Enhancement of CREB, NRF2, and Reduction of AMPKα in SCA17 Cell Models

Figure 6

AMPKα as a therapeutic target in LM-031-treated TBP/Q79-GFP-expressing SH-SY5Y cells. (a) Experimental flow chart. TBP/Q79-GFP SH-SY5Y cells were plated with retinoic acid (10 μM) added on day 1. The next day, LM-031 or licochalcone A (100 nM) was added to the cells. At 4 h postcompound treatment, AICAR (0.1 mM) was added to the cells for 4 h followed by inducing TBP/Q79-GFP expression (Dox, 5 μg/mL) for 6 days. On day 9, the cells were collected for AMPKα and pAMPKα protein analysis (by Western blot, GAPDH as a loading control) or stained with Hoechst 33342 for aggregation and neurite outgrowth analyses (by HCA). (b) Western blot analysis of AMPKα and pAMPKα protein levels in LM-031 or licochalcone A-treated cells treated with AICAR. To normalize, the relative AMPKα or pAMPKα level of AICAR-untreated cells without inducing TBP/Q79-GFP expression was set as 100%. values: comparisons between induced versus uninduced cells (##), compound-treated versus untreated cells (, ), or AICAR-treated versus untreated cells (&) (). (c) Aggregation and (d) neurite outgrowth assays of LM-031 or licochalcone A-treated TBP/Q79-GFP SH-SY5Y cells treated with AICAR. values: comparisons between compound-treated versus untreated cells () or AICAR-treated versus untreated cells (&, &&) (). (e) Representative microscopy images of TBP/Q79-GFP-expressing SH-SY5Y cells treated with LM-031 or licochalcone A, and AICAR-treated cells treated with LM-031 or licochalcone A. Aggregates were marked in red (left) and neurites and cell bodies were outlined (right) for outgrowth quantification.

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