Research Paper Advance Articles pp 23619—23646

Pathomechanism characterization and potential therapeutics identification for SCA3 targeting neuroinflammation

Figure 3. Effects of the tested compounds on cell viability, caspase 1 activity, polyQ aggregation, and neurite outgrowth in ATXN3/Q75-GFP SH-SY5Y cells. (A) Experimental flow chart. ATXN3/Q75-GFP cells were plated on dishes with retinoic acid (RA, 10 μM) added on day 1 to initiate neuronal differentiation. Next day, compound (10 μM) was added to the cells for 8 h followed by inducing ATXN3/Q75-GFP expression with doxycycline or not (± Dox, 5 μg/ml) for 6 days. Cell viability, caspase 1 activity, aggregation and neurite outgrowth were assessed. (B) Relative cell viability and caspase 1 activity (n = 3). For normalization, the relative viability and caspase 1 activity of uninduced and untreated cells was set as 100%. (C) High content polyQ aggregation analysis of ATXN3/Q75-GFP-expressing cells with compound treatment (n = 3). Shown below were filter trap assay of SDS-insoluble ATXN3/Q75-GFP aggregate and Western blot analysis of soluble ATXN3/Q75-GFP protein with compound treatment using GFP antibody (n = 3). To normalize, the relative trapped or soluble ATXN3/Q75-GFP without compound addition was set as 100%. (D) Neurite length, process or branch of ATXN3/Q75-GFP-non-expressing or expressing cells with compound treatment (n = 3).