Fig. 5

Complement C3 released by tau PFF-treated PAs is a primary neurotoxic factor. (A) Immunoblotting analysis of the indicated secretory proteins in conditioned medium from primary astrocytes exposed to tau-PFF, OPN, or PBS. The blots represent three biological repeats. (B-E) The mRNA expression of the indicated genes from the RNAseq experiment in Fig. 1. Error bars indicate means ± SD. *, p < 0.05; **, p < 0.01, ****, p < 0.0001 by one-way ANOVA. n = 3 biological repeats. (F, G) iNeuron viability test after treatment with condition medium (CM) from tau PFF-treated wild type control (Ctrl.) astrocytes or astrocytes with the indicated gene knockdown (KD). The graph in (G) show the quantification of cell death. Error bars indicate means ± SD. *, **, p < 0.01, ****, p < 0.0001 by one-way ANOVA. n = 3 biological repeats. (H) Immunoblotting analysis of the indicated proteins in the condition medium from wild type primary astrocytes or C3- or MPP3-knockdown astrocytes treated with tau PFF. Shown is a representative gel from 3 biological repeats. (I) Neurotoxicity testing after iNeurons were treated with recombinant protein MMP3 or C3 at the indicated concentrations. Cells treated with PBS or tau PFF in parallel were used as negative and positive controls, respectively. (J) Quantification of cell death in (I). Error bars indicate means ± SD, n = 3 biological repeats