Fig. 6: ApoE in non-exosome ACM fractions inhibits A-Exo.-mediated stimulation on neuronal axon growth. | Nature Communications

Fig. 6: ApoE in non-exosome ACM fractions inhibits A-Exo.-mediated stimulation on neuronal axon growth.

From: Astroglial exosome HepaCAM signaling and ApoE antagonization coordinates early postnatal cortical pyramidal neuronal axon growth and dendritic spine formation

Fig. 6

Representative images (a) and quantification (b) of βIII-tubulin+ neuronal axon (white arrows) length following treatment of cortical neurons with A-Exo. or A-Exo. mixed with flowthrough (FT) from the SEC column; 0.2x and 0.5x FT each is concentrated from 2- or 5-ml exosome-free ACM, respectively. Number of neurons quantified in each group shown in the graph (7–9 neurons/replicate, 3 biological replicates)/group; Scale bar: 100 μm; c Representative (from >5 replicates) immunoblot of different apolipoproteins in all eluted fractions (500 μl/fraction, pooled as indicated) of ACM (100 ml) from SEC with optimal exposure. Unconcentrated elution (15 μl/sample) was run on immunoblot; Representative images (d) and quantification (e) of βIII-tubulin+ neuronal axon (white arrows) length following co-treatment of cortical neurons with A-Exo. and different dose of hAPOE3. Number of neurons quantified in each group shown in the graph (4–5 neurons/replicate, 3 biological replicates)/group; Scale bar: 100 μm; f Quantification of βIII-tubulin+ neuronal axon length following co-treatment of A-Exo. with common hAPOE isoforms. Number of neurons quantified in each group shown in the graph (10–12 neurons/replicate, 2 biological replicates)/group; Representative images (g) and quantification (h) of βIII-tubulin+ neuronal axon (white arrows) length in control cortical neurons (i) or neurons treated with A-Exo. (ii) and A-Exo. mixed with WT (iii) or ApoE KO (iv) FT, respectively. Scale bar: 100 μm; Number of neurons quantified in each group shown in the graph (7–9 neurons/replicate, 3 biological replicates)/group; Representative images (i) and quantification (j) of βIII-tubulin+ neuronal axon (white arrows) length in control (i) cortical neurons or neurons treated with WT (ii) or ApoE KO (iii) A-Exo. Number of neurons quantified in each group shown in the graph (4–8 neurons/replicate, 3 biological replicates)/group; Scale bar: 100 μm; 1 μg A-Exo. was used in each treatment. p values in (b, e, f, h, j) were calculated using one-way ANOVA followed by a Tukey post hoc test; n.s. not significant. Data are presented as mean values ± SEM.

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