Fig. 2.

Different conformations of thtt amyloids show distinct cytotoxicity in neuro2a cells. (A) Significant acceleration of thttQ150-GFP aggregation by introduction of in vitro thttQ42 amyloids. Buffer alone (Upper) or in vitro thttQ42 amyloids (Lower) were introduced into thttQ150-GFP stable neuro2a cells. Shown are fluorescent (Left) and DIC (Right) images of the thttQ150-GFP cells after 15 h of thttQ150-GFP expression and cell differentiation. (Scale bar, 250 μm.) (B) Buffer alone, GSTthttQ42 monomer, thttQ42 amyloids, GSTthttQ62 monomer, thttQ62 amyloids, or BSA aggregates were introduced into stable thttQ150-GFP neuro2a cells. The number of cells with thttQ150-GFP foci was counted after 15 h of the thttQ150-GFP expression and cell differentiation. Values are mean ± SD. (C) Thermal stability of the thttQ60-GFP amyloids formed in the presence of in vitro thttQ42-4 °C (thttQ60-GFP[thttQ42-4 °C]) or thttQ42-37 °C amyloid “seeds” (thttQ60-GFP[thttQ42-37 °C]). (D) The band intensity of thttQ60-GFP[thttQ42-4 °C] (blue) or thttQ60-GFP[thttQ42-37 °C] (red) amyloids in C was plotted against temperature. (E, F) Buffer alone, in vitro thttQ42-4 °C amyloids, thttQ42-37 °C amyloids, GSTthttQ42 monomer, or BSA aggregates were introduced into thttQ16-, Q60-, or Q150-GFP neuro2a cells under a regulatable promoter. The thtt expression and cell differentiation started after 3 h of the in vitro amyloid transduction. The number of cells with thtt-GFP foci was counted after 15 h (E) and cell viability was examined by MTT assay after 4 days of the thtt expression and cell differentiation (F). *, P < 0.01. Values are mean ± SD.